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Historic,  Archive  Document 

Do  not  assume  content  reflects  current 
scientific  knowledge,  policies,  or  practices. 


Newsletter  for  the  USDA  Plant  Genome  Research  Program 


Volume  4,  No.  1/2 


USD  As  Office  of  Agricultural 

Biotechnology 

Jean  A.  Larson , M.A. 

Office  of  Agricultural  Biotechnology,  USDA 
Washington,  D.C. 

Can  bioengineered  organisms  of  agricultural  importance 
be  released  safely  into  the  environment  and  the  market- 
place? Will  consumers'  desires  for  labeling  of  biotech 
products  impact  on  the  proposed  Food  and  Drug 
Administration's  policies?  What  will  be  the  impact  of 
BST  on  the  dairy  industry?  Will  biotechnology  technolo- 
gies help  revitalize  rural  America?  Can  this  new 
technology  be  useful  in  preventing  and  detecting 
food  safety  problems?  These  and  other  questions 
are  routinely  being  addressed  by  the  USDA 
Office  of  Agricultural  Biotechnology. 

The  Office  of  Agricultural  Biotechnol- 
ogy (OAB)  was  established  in  1986  by  a 
Secretary's  Memorandum  1020-27.  Its 
role  is  to  coordinate  the  development 
of  consistent  biotechnology  pohcies 
and  procedures  within  USDA. 

Current  OAB  functions  include: 

• Under  a Presidential  Initiative, 

OAB  is  the  USDA  action  office  for  a multi-year 
Federal  initiative  on  biotechnology  research. 

• OAB  staffs  a Federal  advisory  committee,  the 

Agricultural  Biotechnology  Research  Advisory 
Committee  (ABRAC).  This  Committee  pro- 
vides a public  forum  for  issues  in  agricultural 
biotechnology. 

• OAB  staffs  the  Committee  on  Biotechnology  in 

Agriculture  (CBA),  composed  of  six  USDA 
Agency  Administrators  and  two  Assistant 


July  1993-July  1994 


Secretaries,  and  the  Biotechnology  Council, 
composed  of  senior  agency  staff. 

OAB  has  provided  leadership  for  the  development  of 
-biotechnology  guidelines  for  agricultural  research; 
-scientific  exchanges  involving  biotechnology; 
-environmental  assessments  for  transgenic  fish; 
-performance  standards  for  research  with  transgenic 

fish  and  shellfish; 

-advice  when  requested  by  regulatory  agencies; 

-a  biotechnology  consumer  information  plan  for 
USDA; 

-coordinated  responses  on  regulatory  and 
research  issues  to  other  Departments; 
-international  conferences /workshops  on 
animal  and  plant  biotechnology,  includ- 
ing three  international  conferences  on 
The  Biosafety  Results  of  Field  Tests 
of  Genetically  Modified  Plants  and 
Microorganisms"; 

-staff  papers  and  speeches  for  the  Office  of  the  Secretary. 

Since  the  implementation  of  the  January  31,  1992, 
Presidential  Initiative  on  Biotechnology  Research,  OAB 
has  been  the  action  office  for  USDA.  Twelve  Federal 
agencies  participate  in  the  activity.  Their  efforts  on  the 
biotechnology  crosscut  are  coordinated  by  the  Biotech- 
nology Research  Subcommittee  (BRS)  of  the  Committee 
on  Fundamental  Science  and  Engineering  Research  and 
Development  of  the  National  Science  and  Technology 
Council.  As  a participant,  OAB  has  collected  research 
program  and  budget  data  from  the  Agricultural  Research 


2 


Probe 


Volume  4,  No.1/2 


Service,  Cooperative  State  Research 
Service,  Economic  Research  Service 
and  the  Forest  Service  and  eight  non- 
USDA  agencies  which  are  involved 
in  agriculturally  related  programs 
and  assembled  data  into  a consistent 
format  for  reporting. 

The  ABRAC  consists  of  15 
experts  from  academia,  industry, 
government,  and  public  interest 
groups  with  knowledge  and  experi- 
ence in  one  or  more  of  the  following 
areas:  recombinant  DNA  research  in 
plants,  animals,  and  microbes; 
ecology  and  environmental  science; 
agricultural  production  practices; 
biological  containment  and  field 
release;  applicable  laws  and  regula- 
tions; standards  of  professional 
conduct  and  practice;  public  atti- 
tudes; public  health /epidemiology; 
and  occupational  health  and  ethics. 
Fifteen  ABRAC  members  have  been 
recently  appointed  by  the  Secretary 
of  Agriculture. 

The  purpose  of  ABRAC  is  to 
advise  the  Department,  through  the 
Assistant  Secretary  for  Science  and 
Education,  with  respect  to  policies, 
programs,  operations,  and  activities 
associated  with  questions  of  biosafety, 
the  development  of  guidelines  and 
performance  standards  for  research 
with  genetically  modified  organisms, 
and,  in  response  to  a specific  re- 
quest, the  development  of  recom- 
mendation for  the  food  safety 
evaluation  of  transgenic  livestock. 

The  most  important  issue  that 
will  be  addressed  by  the  new  Com- 
mittee in  the  coming  months  will  be 
to  complete  the  development  of 
performance  standards  for  outdoor 
research  with  genetically  modified  fish 
and  shellfish.  Other  important  issues 


may  include:  management  of  resis- 
tance to  biopesticides  in  crop  plants; 
production  of  pharmaceuticals  in 
plants  and  animals;  use  and  effects  of 
synthetic  sequences  in  organisms  of 
agricultural  importance;  risk  man- 
agement and  risk  communications; 
and  public  attitudes,  perceptions, 
and  acceptance  of  genetically  engi- 
neered products. 

In  addition  to  the  scientific 
aspects  of  modern  biotechnology, 

OAB  clearly  recognizes  the  public 
relations  dimension  of  biotechnology. 
A well-informed  public  is  better  able 
to  participate  in  the  decision-making 
process  about  biotechnology.  Toward 
this  end,  OAB  shares  information  with 
the  media,  participates  in  public 
affairs  activities  around  the  country, 
and  contributes  articles  on  biotechnol- 
ogy to  the  general  as  well  as  the 
scientific  press.  The  Office  publishes  a 
monthly  newsletter.  Biotechnology 
Notes.  The  newsletter  highlights 
activities  on  biotechnology  issues  at 
USDA  and  in  the  private  sector.  The 
dissemination  of  the  newsletter  is  via 


mail,  through  USDA's  Computerized 
Information  Delivery  System,  and  on 
Internet  through  the  National  Agri- 
cultural Library's  Gopher;  provided 
by  the  Biotechnology  Information 
Center.  From  time  to  time  the  Office 
has  sponsored  national  and  interna- 
tional conferences  and  workshops  on 
biotechnology  topics. 

The  international  and  trade 
implications  of  agricultural  biotech- 
nology are  of  growing  concern  to 
many  U.S.  economic  planners  and 
policymakers.  On  the  international 
front,  OAB  is  involved  in  studying 
research  and  technology  transfer 
programs  in  competing  nations; 
promoting  international  consensus  on 
the  scientific  principles  that  underlie 
the  environmental  and  human  safety 
of  agricultural  biotechnology;  and 
working  with  the  U.S.  Trade  Repre- 
sentative, the  Food  and  Drug  Admin- 
istration and  the  Foreign  Agricultural 
Service  to  provide  information  to 
trading  partners  regarding  U.S.  food 
safety  procedures. 


Table  of  Contents 

USDA's  Office  of  Agricultural  Biotechnology 1 

Summary  of  the  1993  Plant  Genome  Awards  4 

RBNET  (Electronic  Mail  Network  for  Rice  Biotechnologists) 7 

New  Capabilities  and  Connections  for  the  Plant  Genome  Database  9 

Plant  Genome  II  Conference  Report 22 

Soybase  News  24 

The  Class  of  1993  Plant  Genome  Grant  Recipients  25 

A Primer  on  Images  and  the  Internet 20 

Maize  Genome  Database,  a USDA-ARS  Plant  Genome  Database  22 

Introducing  Dr.  Edward  Kaleikau  24 

Announcing  Plant  Genome  III  Meeting  25 

Calendar  of  Upcoming  Genome  Events  27 

Survey  of  Synonymous  Codon  Usage  in  Nuclear  Genes  of  Arabidopsis,  Soybean  and  Maize  .30 

Plant  Genome  Analysis  by  Single  Arbitrary  Primer  Amplification  32 

Distinctive  Biology  of  Forest  Trees  Highlighted  at  Sixth  International  Meeting  37 


July  1993  -July  1994 


Probe 


3 


Agricultural  Biotechnology  Research  Advisory  Committee 


Dr.  Walter  A.  Hill 

School  of  Agri.  & Home  Economics 

Tuskegee  University 

Dr.  Anne  R.  Kapuscinski 
Dept,  of  Fisheries  and  Wildlife 
University  of  Minnesota 

Dr.  Pamela  G.  Marrone 
Entotech,  Inc. 


Dr.  Deborah  K.  Letourneau 
Board  of  Environmental  Studies 
Univ.  of  California 

Dr.  Rudy  Wodzinski 
Dept,  of  Molecular  Biology  and 
Microbiology 
Univ.  of  Central  Florida 

Dr.  Roy  Fuchs 

Monsanto  Agricultural  Company 


Dr.  James  Tiedje 

NSF  Center  for  Microbial  Ecology 
Michigan  State  Univ. 


Dr.  Ronald  R.  Sederoff 
Dept,  of  Forestry 
North  Carolina  State  Univ. 

Dr.  James  Lauderdale 
The  Upjohn  Company 


Dr.  Susan  Harlander 
Director,  Dairy  Foods 
Land  O'Lakes,  Inc. 

Dr.  Stanley  Pierce 
Rivkin,  Radler,  Bayh,  Hart,  & 
Kremer 

Dr.  Fernando  Osorio 
Department  of  Veterinary 
Biomedical  Sciences 
Univ.  of  Nebraska 

Dr.  H.  Alan  Wood 
Boyce  Thompson  Inst,  for  Plant 
Research 

Dr.  Walter  Reid 
World  Resources  Institute 


Dr.  Paul  Thompson 

Center  for  Biotechnology  Policy  and  Ethics 
Texas  A&M  Univ. 

Minutes  from  ABRAC  meeting  are  published  and  available  from  the  OAB  on  request. 
(703)  235-4419  _ 


Needless  to  say,  the  OAB 
program  is  a very  dynamic  office  that 
maintains  timely  responsiveness  to  the 
ever-changing  biotechnology  scene. 

All  the  above  activities  are  done 
with  a small  core  of  permanent  staff, 
but  OAB  Director  Dr.  Alvin  Young 
says  that  "much  of  my  staff  work  is 
done  by  individuals  on  temporary 
assignment  to  OAB  from  other  USDA 
agencies."  Specialists  in  agricultural 
research,  extension /technology 


transfer,  regulations,  environmental 
impact,  economics,  public  relations,  and 
international  affairs  have  been  supplied 
to  OAB  by  cooperating  agencies.  At  the 
completion  of  their  assignments,  says 
Young,  "these  individuals  take  their 
new  biotechnology  knowledge  and 
experience  with  them  back  to  their 
agencies  and  everyone  benefits." 

For  additional  information  or  to 
request  OAB  published  materials, 
contact  the  OAB  at  (703)  235-4419.  ♦ 


Probe 


ISSN:  1057-2600 

The  official  quarterly  publication  of 
the  USDA  Plant  Genome  Research 
Program.  This  newsletter  is  aimed 
at  facilitating  interaction  throughout 
the  plant  genome  mapping  commu- 
nity and  beyond. 

Probe  is  a publication  of  the  Plant 
Genome  Data  and  Information 
Center,  National  Agricultural 
Library. 

Managing  Editor 
Susan  McCarthy,  Ph.D. 

Editor 

Joanne  Meil 

Production  Manager 
Terrance  Henrichs 

Layout  and  Design 
Terrance  Henrichs 

Special  Thanks  to: 

Barbara  Buchanan 
Andrew  Kalinski 
Stephanie  MeGee 
Marcia  Norfleet 

Articles,  announcements,  and 
suggestions  are  welcome. 

Correspondence  Address 
Susan  McCarthy,  Ph.D. 

NAL,  4th  Floor 

10301  Baltimore  Blvd 

Beltsville,  MD  20705 

Phone:(301)504-6613 

FAX:  (301)504-7098 

email:  smccarth@.nalusda.gov 

USDA  Program  Office 
Dr.  Jerome  Miksche 
USDA/ ARS/NPS/PNRS 
Room  33 1C,  Bldg  005 
BARC-WEST 
Beltsville,  MD  20705 
Phone:(301)504-6029 
FAX:  (301)504-6231 


National  Agricultural  Library 


Probe 


Volume  4,  No.1/2 


Competitive  Edge 


$ 

Summary  of  the  1993  Plant  Genome 

Awards 


Dr.  Ed  Kaleikau,  Program  Director 
National  Research  Initiative  Competitive  Grants  Program 
Cooperative  State  Research  Service,  USDA 
Washington,  DC  20250 


Table  1 


7n  1993,  Congress  appropri- 
ated $97.5  million  for  the 
National  Research  Initiative 
Competitive  Grants  Pro- 
gram (NRICGP),  of  which  $12.1 
million  was  made  available  for  plant 
genome  research.  The  Plants  Divi- 
sion of  the  NRICGP  in  USDA's 
Cooperative  State  Research  Service 
administers  the  plant  genome 
grants.  In  addition  to  the  NRICGP 
allocation,  $3.67  million  was  appro- 
priated to  USDA's  Agricultural 
Research  Service  (ARS)  for  program 
management,  setting  targets  for 
genome  mapping  research,  and 
database  development  for  agricul- 
turally important  plants.  After 
program  administration  costs,  the 
net  amount  available  for  all  plant 
genome  research  totalled  $15.1 
million:  $3.0  million  for  the  ARS 
and  $12.1  million  for  the  NRICGP. 
Mission-oriented  research  proposals 
that  address  the  goal  of  improving 
agronomic  qualities  through  ge- 
nomic research  were  submitted  to 
the  NRICGP  by  scientists  from  the 
research  community.  Each  proposal 
was  peer-reviewed  by  experts  in  the 
area  of  genomic  research  and  was 

Continued  page  7 


SPECIES  REPRESENTED  BY  PLANT  GENOME  AWARDS  1993 

NUMBER  OF 

DOLLAR 

SPECIES 

AWARDS 

AMOUNT 

Agrobacterium 

2 

250,000 

Alfalfa 

2 

220,000 

Apple 

1 

276,000 

Arnbidopsis 

8 

539,000 

Barley 

5 

961,000 

Bean 

2 

370,000 

Blueberry 

1 

118,422 

Brassica 

4 

262,000 

Cotton 

2 

299,013 

Douglas  Fir 

1 

223,000 

Lettuce 

1 

200,000 

Maize 

20 

2,405,700 

Millet 

1 

120,000 

Mungbean 

1 

115,000 

Oats 

2 

300,000 

Peach 

1 

200,000 

Peanut 

2 

262,000 

Petunia 

2 

211,000 

Pine 

3 

453,000 

Poplar 

1 

120,000 

Potato 

2 

190,000 

Rice 

3 

570,000 

Sorghum 

4 

297,200 

Soybean 

3 

500,000 

Tobacco 

11 

1,392,400 

Tomato 

7 

846,900 

Wheat 

6 

1,145,200 

Wild  Rice 

i 

40,000 

Totals 

91 

12,126,238 

ARS  G.A.  Smith,  J Miksche 


* Summations  of  award  numbers  and  dollar  amounts  do  not 
sum  to  the  actual  values  presented  because  some  of  the  awards 
utilized  more  than  one  species. 


July  1993  -July  1994 


Probe 


5 


Table  2 

GENE  SYSTEMS  OF  TRAITS,  NRI  PLANT  GENOME  GRANTS,  1993 


AC /DC  transposons 

AC/Ds  mutagenesis  and  integration 

Alcohol  dehydrogenase 

Amylase  activation 

Apomixis 

Bacterial  blight  resistance 

Ca+  Modulated  leaf  receptor  protein 

Cytokinin  response 

Disease  resistance 

Drought  tolerance 

Floral  homeotic  genes  and  sterility 

Flowering 

Gene  targeting  for  excision  of  foreign  DNA 
Lipid  desaturation 
mRNA  stability 

Phytochrome  A mRNA  degradation 

Plasmid  directed  conjugation 

Polyamines  and  stress  tolerance 

QTLs  for  wood  quality 

QTLs  for  yield 

Ribosomal  protein  synthesis 

Rust  resistance 

Seed  maturation 

Stable  transformation 

Starch  synthesis 

Targeted  DNA  integration 

Trichomes  and  insect  resistance 

T-DNA  transport  and  integration 


Anthocyanin  biosynthesis 
Aphid  resistance 
Blight  resistance 

Endodormancy  chilling  requirement 

Ethylene  biosynthesis 

Fertility 

Fiber  quality 

Fruit  quality 

Hessian  fly  resistance 

Inflorescence  development 

Kernel  starch 

Kernel  sucrose  metabolism 
Leaf  epidermal  growth 
Mildew  resistance 
Mitochondria  protein  synthesis 
Nematode  resistance 
Nodulation  and  N fixation 
Phaseolin  and  seed  protein 
Photorespiration 
Plastid  light  response 
Rust  resistance 
Seed  oil  synthesis 
Self  incompatibility 
Ubiquitin  ligation 
Vigor  and  plant  morphology 
Virulence  genes 
Virus  resistance 
Wood  specific  gravity 


6 


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Volume  4,  No.1/2 


Table  3 


GENETIC  PHENOMENA  NRI  PLANT  GENOME  GRANTS  1993 

DESCRIPTION 

SPECIES 

Amylase  activation  & repression 

Barley,  Maize 

Annual  growth  rate 

Douglas  fir 

Anthocyanin  biosynthesis 

Maize 

Apomixis,  asexual  reproduction 

Pearl  millet 

Cell  growth 

Tobacco 

Chromosome  recombination 

Wheat 

Chromosome  sorting  libraries 

Tomato 

Cloning  disease  resistance  genes 

Rice 

Cytokinin  response 

Arabidopsis 

Developmental  regulation 

Tomato,  Tobacco 

Drought  tolerance 

Grain  sorghum 

Edthylene  biosynthesis 

Tomato 

Endodormancy  & chilling  requirement 

Blueberry 

Gene  effects  on  chromatin  structure 

Maize 

Gene  tagging  for  insect  resistance 

Maize 

Genetic  engineered  sterility 

Poplar 

Gene /chromosome  identification 

Wheat 

Homologous  recombination 

Arabidopsis 

Homozygous  deletion  stocks 

Wheat 

Hormonal  control  of  seed  maturation 

Tobacco 

Inflouresence  development,  flowering 

Maize,  Tobacco 

Lipid  desaturation 

Peanut 

Mapping  cold,  disease  resistance 

Peach 

mRNA  stability  in  dicots 

Tobacco 

Mutation  via  transposable  elements 

Tomato 

Nodulation  & N fixation 

Bean,  Medicago 

Nuclear  targeting  of  DNA 

Agrobacterium 

Nuclear  & organelle  DNA  variation 

Barley 

Nuclear-plastid  communication 

Tobacco 

Photorespiration,  nitrogen  assimilation 

Ambidopsis 

Phytochrome  A,  mRNA  degradation 

Oats 

Plasmid  directed  conjugation 

Agrobacterium 

Phytochrome  gene  control 

Tomato,  Rice,  Sorghum 

Repeat  induced  gene  silencing 

Arabidopsis 

RNA  editing 

Petunia 

Seed  oil  synthesis 

Maize,  Brassica 

Self  incompatibility 

Brassica 

Sequence  repeats,  microsatellites 

Soybean 

Site  directed  mutagenesis 

Maize 

Spatial  organization  of  genome 

Maize,  Sorghum 

Starch  metabolism  & transport 

Maize 

Tandem  repeat  sequences 

Rice 

Targeted  DNA  integration 

Tobacco 

Transformation  disease  resistance 

Potato 

Transformation  drought  tolerance 

Pine 

Transformation  insect  resistance 

Tomato 

Transformation  virus  resistance 

Peanut 

Transposable  elements 

Maize 

Transposon  tagging 

Maize 

T-DNA  transfer,  and  transport 

Tobacco,  Maize 

Ubiquitin  activating  enzymes 

Arabidopsis 

Wound  inducible  insect  resistance 

Tobacco 

Probe 


July  1993 -July  1994 


Awards— Continued  from  page  4 

judged  on  its  scientific  and  technical 
merit,  qualifications  of  proposed 
personnel,  and  relevance  to 
sustainability  and  stated  research 
objectives  in  the  solicitation  for 
proposals. 

In  1993,  the  majority  of  plant 
genome  funds  supported  awards 

Table  4 


made  in  two  programs  in  the 
NRICGP  Plant  Division:  the  Plant 
Genome  Program  and  the  Plant 
Genetic  Mechanisms  Program.  A small 
portion  of  funds  also  supported 
genome-related  research  in  other 
progams  of  NRICGP.  Research  projects 
being  supported  in  FY  1993  are  summa- 
rized in  tire  accompanying  tables. 


TECHNIQUE  DEVELOPMENT  NRI  PLANT  GENOME  GRANTS,  1993 


Antibody  synthesis  for  proline 

Biolistic  transformation  with  high  molecular  weight  DNA 
Chromosome  painting 

Construction  and  insertion  of  chimeric  gene 

Develop  target  probes  for  fly  resistance 

DNA  vector  synthesis 

Functional  cloning  of  disease  resistant  genes 

Identification  of  effective  sterility  genes 

Isolate  nuclear  skeletal  structures 

Map  based  cloning  system  for  Rice 

Megabase  DNA  isolation 

Molecular  probe  and  reagent  development 

Regeneration  after  transformation  for  Pine 

Substrate  features  required  for  enzyme  activation 

Transfer  teosinte  chromosomes  to  maize 

Transformation  system  for  Peanut 

Transposon  tagging 

Wound  induced  transformation 


Off  the  Wire 


Ninety-one  awards  were  made  to 
scientists  from  34  States  (See  article, 
"The  Class  of  1993  Plant  Genome 
Grant  Recipients"  in  this  issue). 

Nineteen  agronomic,  horticul- 
tural, and  forest  tree  species  are 
undergoing  genetic  and  physical 
mapping  procedures  (Table  1).  Fifty- 
six  genes  and  gene  systems  are  being 
studied  (Table  2),  as  well  as  genetic 
phenomena  in  various  plant  species 
(Table  3).  Many  new  molecular 
techniques  are  being  pursued  (Table 
4).  The  data  were  supplied  by  the 
NRICGP  staff,  and  compiled  into 
tables  by  Dr.  G.S.  Smith  and  Dr.  J.P. 
Miksche  in  ARS.  ♦ 


Plant  Genome 
Grant 

Deadline  Dates 
Announced 

See  page  8 
in  this  issue 


RBNET  (Electronic  Mail  Network  for 

Rice  Biotechnologists) 


With  support  from  the  Rockefeller 
Foundation,  an  electronic  mail 
network  has  been  established  to 
increase  communication  among  the 
international  rice  biotechnology 
community.  This  network  (RBNET) 
is  being  managed  by  Professor 
Verma  at  the  Ohio  State  Biotechnol- 


ogy Center,  1060  Carmack  Road, 
Columbus,  Ohio  43210  USA.  To 
address  all  users  of  the  network  you 
may  send  a message  to: 
rbnet@magnus.acs.ohio-state.edu.  To 
add  your  name  to  the  RBNET 
mailing  list,  please  send  your  E-Mail 
address  to: 

dverma@magnus.acs.ohio-state.edu. 


For  those  colleagues  in  coun- 
tries where  electronic  mail  service  is 
not  currently  available,  attempts  are 
being  made  to  deliver  a message  by 
fax.  If  any  one  of  you  are  in  this 
situation  and  wish  to  communicate 
to  the  RBNET  users,  you  may  fax 
your  message  to  the  attention  of 
Professor  Verma  at  (614)  292-5379 
and  your  message  will  be  posted  on 
the  RBNET  for  distribution.  ♦ 


Probe 


Volume  4,  No.1/2 


1995  NRI  Grant  Deadlines 


Postmarked 

Dates 

Program 

Codes 

Program 

Areas 

Contacts 

(202) 

November  14, 1994 

31.0 

Improving  Human  Nutrition  for  Optimal  Health 

205-0250 

52.1 

Plant  Genome 

401-1901 

52.2 

Plant  Genetic  Mechanisms 

401-5042 

November  21,  1994 

22.1 

Plant  Responses  to  the  Environment 

401-4871 

December  5, 1994 

51.1 

Plant  Pathology 

401-4310 

54.1 

Photosynthesis  and  Respiration 

401-6030 

December  12,  1994 

25.0 

Soils  and  Soil  Biology 

401-4082 

December  19,  1994 

26.0 

Water  Resources  Assessment  and  Protection 

401 -4504 

51.5 

Biological  Control  Research 

401-5114 

January  9,  1995 

51.2 

Entomology 

401-5114 

51.3 

Nematology 

401-5114 

51.4 

Weed  Science 

401-4310 

44.0 

Sustaining  Animal  Health  and  Well-Being 

401  -6303 

January  17,  1995 

71.1 

Food  Characterization/Process/Product  Research 

401-1952 

71.2 

Non-Food  Characterization/Process/Product  Research 

401-1952 

72.0 

Biofuels  Research 

401-1952 

41.0 

Enhancing  Animal  Reproductive  Efficiency 

401  -6234 

January  23,  1995 

23.0 

Forest/Range/Crop/Aquatic  Ecosystems 

401-4082 

53.0 

Plant  Growth  and  Development 

401-5042 

January  30,  1995 

32.0 

Ensuring  Food  Safety 

401-4399 

51.6 

Assessing  Pest  Control  Strategies 

401-5114 

100 

Agricultural  Systems 

401-1901 

401-6303 

February  6,  1995 

54.2 

Nitrogen  Fixation/Nitrogen  Metabolism 

401-6030 

61.0 

Markets  and  Trade 

401-3487 

62.0 

Rural  Development 

401-3487 

February  13, 1995 

73.0 

Improved  Utilization  of  Wood  and  Wood  Fiber 

401-4871 

February  21,  1995 

42.0 

Improving  Animal  Growth  and  Development 

205-0250 

43.0 

Identifying  Animal  Genetic  Mechanisms  and  Gene 

Mapping 

401-4399 

February  27,  1995 

80.1 

Research  Career  Enhancement  Awards 

401-6234 

80.2 

Equipment  Grants 

401-6234 

80.3 

Seed  Grants 

401-6234 

The  1995  program  solicitation  and 
application  kit  will  be  available  in 
September.  Please  note  that  potential 
applicants  who  are  on  the  competi- 
tive research  grants  mailing  list,  who 
sent  in  applications  in  fiscal  year 
1994,  or  who  recently  requested 
placement  on  the  list  for  fiscal  year 
1995  will  automatically  receive 
copies  of  the  program  solicitation 


and  the  NRICGP  application  kit.  All 
others  may  request  copies  from: 
National  Research  Initiative  Com- 
petitive Grants  Program  (NRICGP) 
c/o  Proposal  Services  Branch 
AMD/CSRS/USDA 
AG  Box  2245 

Washington,  DC  20250-2245 

Telephone:  (202)  401-5048 

For  more  information  about  the  Plant 


Genome  Grants  Program, 
contact: 

Dr.  Ed  Kaleikau 
Program  Director 
NRICGP /CSRS/ USD  A 
AG  Box  2241 

Washington,  DC  20250-2241 
Telephone:  (202)  401-1901  ♦ 


Probe 


July  1993  -July  1994 

9 

Home  Base 

i 

I 

i 

New  Capabilities  and  Connections  for 

the  Plant  Genome  Database 


Stephen  M.  Beckstrom-Sternberg 
Plant  Genome  Data  and  Information  Center 
National  Agricultural  Library,  USDA 
Beltsville,  MD  20705-2351 


The  Plant  Genome  Database  has  been 
considerably  augmented  in  the  past 
few  months.  A number  of  new 
connections  and  capabilities  have 
been  added  to  the  World  Wide  Web 
(WWW)  and  gopher  interfaces,  and  a 
text-based  Lynx  client  has  been 
added  to  provide  access  to  those 
with  slower  Internet  connections 
and/or  low  resolution  graphics.  The 
following  is  a summary  list  of  recent 
changes  to  the  Plant  Genome  Data- 
base: 

1)  WWW  Interface  to  ACEDB 
Databases,  2)  New  Query  Capabili- 
ties for  Plant  Genome  Data,  3)  Links 
to  External  Data  (Agricola,  GenBank, 
Swiss-Prot,  Images),  4)  New  Gopher 
Services,  5)  Telnet  Access  to  WWW 
Lynx  Client,  6)  Collaborator's  Page 
summarizes  Collaborator's  Services, 
7)  New  Draft  for  Plant  Genome  III 
Meeting,  8)  New  Solanaceae  Data- 
base Shows  Syntenic  Relationships 

WWW  Interface  to  ACEDB 
Databases 

The  Plant  Genome  Database  Project 
is  now  offering  a collection  of 
ACEDB-based  databases  for  both 


plant  and  non-plant  genomes.  The 
databases  have  been  installed  behind 
the  "Moulon"  ACEDB-WWW  server 
and  can  be  browsed  and  queried 
interactively.  The  various  databases 
are  listed  below: 

Plants 

AAtDB — Arabidopsis 
SoyBase — soybeans 
RiceGenes — rice 
MaizeDB — maize 
GrainGenes — wheat,  barley,  rye, 
relatives 

TreeGenes — forest  trees 
SolGenes — Solanaceae 

Other  organisms 
ACeDB — C.  elegans 
AceMap — Human  Chromosome  X 
FlyDB — Drosophila  melanogaster 
MycDB — mycobacteria 
AGsDB — A Genus  species  Database 

New  Query  Capabilities  for 
Plant  Genome  Data 

The  ACEDB-based  databases  can  be 
queried  using  the  ACEDB  query 
language,  query-by-example  (QBE) 
and  a query-building  (QB)  tool.  The 


ACEDB  query  language  requires  an 
understanding  of  syntax,  but  QBE 
and  QB  do  not.  In  addition,  all  the 
data  from  the  ACEDB  databases  has 
been  indexed  for  searches  using 
WAIS.  Finally,  data  can  be  searched 
using  agrep — a tool  which  allows 
you  to  make  approximate  matches 
(aka,  fuzzy  searching). 

Links  to  External  Data: 
AGRICOLA,  GenBank, 
SwissProt,  Images 

HTML  links  are  increasingly  being 
used  to  connect  plant  genome  data 
with  data  from  external  sources,  for 
example  AGRICOLA,  GenBank,  and 
SwissProt.  This  allows  us  to  take 
advantage  of  the  division  of  labor 
between  databases  specializing  in 
different  tasks.  For  example,  the 
receptor  kinase  sequence  object  from 
the  Arabidopsis  database  AAtDB 
contains  a link  to  GenBank  labeled 
"M80238".  Clicking  on  it  will  retrieve 
a GenBank  record  from  a computer 
at  National  Institutes  of  Health. 


Mention  of  a trade  name  or  brand  does  not  constitute 
endorsement  or  recommendation  by  the  Department 
over  similiar  products  not  named 


Probe 


New  Gopher  Services 
The  plant  genome  information  on  the 
agricultural  genome  Gopher  has 
been  reorganized.  It  can  be  used  to 
access  the  same  information  that  is 
presented  by  the  WWW  interface  to 
the  genome  databases,  both  plant 
and  non-plant. 

Telnet  Access  to  WWW  Lynx 
Client 

Users  who  are  restricted  to  VT100  or 
VT200  terminals  or  emulators  can 
access  the  WWW  services  using 
Lynx,  a text-only  client  (see  below  for 
access  details).  The  client  is  set  up  so 
that  only  NAL  data  (and  data  from  a 
few  other  sources)  is  available — it 
cannot  be  used  to  surf  the  Internet 
(sorry). 

Collaborator's  Page 
Summarizes  Collaborator's 
Services 

A page  has  been  set  up  with  hnks  to 
resources  maintained  independently 
by  the  plant  genome  collaborators. 


The  services  include  ftp  archives, 
gophers,  and  access  to  databases. 

New  Draft  for  Plant  Genome 
III  Meeting 

There  is  a new  version  of  a draft 
describing  the  next  plant  genome 
meeting  in  San  Diego  in  mid-January 
1995  (gopher  and  WWW). 

New  Solanaceae  Database 
Shows  Syntenic 
Relationships 

SolGenes,  a database  for  pepper, 
tomato,  and  potato,  now  shows  the 
syntenic  relationships  between 
chromosomes  from  these  members  of 
the  Solanaceae.  The  information  is 
presented  as  an  on-line  image  of  a 
genetic  map.  The  image  responds  to 
mouse  clicks  enabling  one  to  find  out 
more  about  a particular  locus  by 
clicking  on  it,  then  selecting  "Show 
as  text."  Figure  1 is  an  example 
comparing  chromosome  9 of  tomato 
and  potato.  Access  to  these  graphi- 
cally depicted  relationships  from  the 


Volume  4,  No.1/2 


agricultural  genome  home  page  is 
via  the  following  sequential  menu 
selections  (WWW  Interface  to 
ACEDB  Data,  Browse  Solgenes,  and 
Multimap). 

Ways  to  Access  the 
Agricultural  Genome 
Databases 

WWW: 

http:/  /probe.nalusda.gov:8000/ 
Gopher: 

gopher  probe.nalusda.gov 
Lynx: 

telnet  probe.nalusda.gov 
login:  lynx 

password:  (none-hit  return) 

Anonymous  FTP: 
probe.nalusda.gov 

♦ 


Plant  Genome  III  Meeting  Announcement 


The  Science  and  Technology  Coordinating  Committee  of  the  USDA  Plant 
Genome  Research  Program  will  hold  an  open  meeting  on  Monday, 
January  16,  1995  from  7:30  pm  to  10:00  pm.  The  Committee  will  be 
reviewing  the  progress  of  the  Plant  Genome  Research  Program.  All  Plant 
Genome  III  attendees  are  invited  to  participate  and  express  their  views. 


July  1993  -July  1994 


Probe 


11 


Isolation  and  Transformation  of  Agriculturally 
Important  Genes 
Instrumentation /Technology 
Applications  of  cDNA  Research 


Chromosome  Structure 
ALFPs/QLTs/Metabolic  Pathways 


$300  Advance  Registration  up  to  December  1,  1994 
$350  after  December  1,  1994  and  on-site 
$100  $tudent  (Pre-Ph.D)  registration 
(Requires  a letter  of  certification  from  department  chairperson.) 


Darrin  Scherago 
Scherago  International,  Inc. 

11  Penn  Plaza,  Suite  1003 
New  York,  NY  10001 
(212)  643-1750 
FAX  (212)  643-1758 

E-mail:  SCHERAGO@BIOTECHNET.COM 


Probe 


12 


Volume  4,  No.1/2 


Connections 


Plant  Genome  II  Conference  Report 


Susan  McCarthy,  Coordinator 

Plant  Genome  Data  & Information  Center 

National  Agricultural  Library,  USDA 


Beltsville,  MD  20705 

Plant  Genome  II  featured 
applications  of  genome 
mapping  and  analysis  to 
solve  existing  problems 
and  uncover  answers  to  fundamental 
questions  relating  to  the  plant 
genome  and  its  evolution.  The 
meeting  was  held  in  San  Diego, 
California  from  Janaury  24  to  Janu- 
ary 27,  1994,  and  attracted  553 
participants  from  22  countries. 

According  to  Steven  Oliver, 
University  of  Manchester,  Manches- 
ter, United  Kingdom,  we  are  entering 
a stage  where  the  taxonomy  of  gene 
function  will  be  essential  in  effi- 
ciently identifying  new  genes.  This 
new  stage  will  require  a 
multidisciplinary  approach  encour- 
aging the  collaboration  of  physiolo- 
gists, geneticists,  biochemists,  and 
plant  breeders.  He  defined  this  era  as 
a new  voyage  of  the  Beagle. 

The  message  was  reinforced  by 
Dr.  James  Cook,  USDA,  ARS  and 
also  Senior  Scientist  at  CSRS,  who 
pointed  out  that  now  is  the  time  to 
bring  plant  breeders  together  with 
molecular  biologists  to  conduct  a 
gene  hunt  for  agronomically  impor- 
tant genes. 

New  Insights 

Understanding  plant  genome  struc- 
ture and  organization  can  lead  to 


interesting  and  relevant  discoveries, 
as  highlighted  by  Dr.  Richard  Flavell, 
Director,  John  Innes  Institute,  Nor- 
wich, United  Kingdom.  According  to 
Flavell,  understanding  the  role  of 
epigenetic  regulation,  gene  order, 
and  in  situ  homology  sequence 
searching  will  ultimately  help  in  the 
practical  application  of  biotechnol- 
ogy. Plants  have  had  to  defend 
themselves  from  foreign  DNA  over 
the  millennia  and  as  a result  have 
developed  strategies  --  including 
gene  silencing  --  to  cope  with 
transposon  selection  pressures.  The 
plant's  ancient  art  of  anti-sense 
technology  may  take  advantage  of 
gene  location.  Gene  location  would 
determine  epigenetic  DNA  methyla- 
tion  events,  which  in  turn  would 
regulate  gene  expression. 

In  all,  Flavell  points  out, 
concerted  evolution  in  the  long  term 
helps  to  maintain  high  levels  of 
conservation  across  the  chromosome 
both  in  terms  of  sequence  and  gene 
order  or  synteny. 

Thomas  Bureau,  University  of 
Georgia,  detected  evidence  of  ancient 
transposon  and  retrotransposon 
events.  Extensive  sequence  similarity 
searches  were  performed  on  the 
GenBank  and  EMBL  nucleotide 
sequence  databases.  These  "database 
mining"  experiments  have  identified 


over  100  normal  plant  gene  se- 
quences showing  evidence  for  a 
member  of  either  the  "Tourist"  or 
"Stowaway"  family  of  transposons. 
The  location  of  several  elements 
corresponds  to  previously  reported 
cis- acting  regulatory  elements. 
Significantly,  a "Tourist"  element 
was  found  to  serve  as  the  promoter 
for  the  maize  auxin-binding  protein 
(abpl).  The  first  plant 
retrotransposon,  Bsl,  was  found  to 
contain  a cellular  gene  fragment;  this 
provides  the  first  evidence  for 
transduction  by  a retrotransposon  in 
plants. 

Progress  in  Rice 

The  Japanese  Rice  Genome  Program 
reported  significant  advances.  Dr. 
Nori  Kurata,  NIAR/STAFF,  Japan, 
described  a genetic  map  with  1,400 
RFLP  and  RAPD  markers.  Over 
7,500  clones  from  callus  tissue  at 
different  developmental  stages  have 
been  sequenced.  Of  those  sequenced 
1,800  are  clones  of  known  function. 
Dr.  Kurata  reports  that  an  expression 
map  has  been  constructed  using 
cDNA  mapping  as  a base.  This  map 
includes  information  on  tissue- 
specificity,  distribution  of  isozyme 
genes,  gene  families,  and  functionally 
related  genes  in  the  genome,  such  as 
ribosomal  protein  genes  and  the 
histone  gene  family. 

Physical  mapping  in  the  Japa- 
nese Rice  Genome  Program  will  be 
used  to  identify  economically  impor- 
tant genes.  Two  YAC  and  three 


July  1993 -July  1994 


Probe 


13 


cosmid  clone  libraries  have  been 
developed,  representing  about  20% 
of  the  rice  genome.  Ordered  libraries 
will  be  prepared  from  these  clone 
libraries.  To  date,  120  YAC  end- 
clones  have  been  isolated  and  end- 
clone  mapping  is  underway.  Typical 
YACs  have  400  kb  inserts,  which, 
when  finished,  the  Japanese  expect 
will  cover  the  rice  genome  six  times 
over. 

Chromosomes  1,  4,  6,  and  11 
are  being  given  high  priority.  It  is 
known  that  a number  of  important 
resistance  genes  reside  on  Chromo- 
some 6.  Mapping  data  from  the 
Japanese  program  have  been  entered 
onto  two  versions  of  an  internal 
database  called  RiceBase.  One 
version  contains  mostly  cDNA 
information,  while  the  other  version 
has  the  physical  map  data. 

International  collaboration  of 
rice  mapping  efforts  was  encouraged 
by  an  informal  workshop  held  in 
conjunction  with  the  conference.  Dr. 
Susan  McCouch,  Cornell  University, 
and  Dr.  Gou-fan  Hong,  Director, 
Chinese  Rice  Genome  Program,  co- 
chaired the  session.  Highlights 
included  Dr.  Kurata's  announcement 
that  the  Japanese  mapping  data 
should  be  made  public  later  this 
year.  Five  prime  sequence  data  for 
several  hundred  markers  are  cur- 
rently available.  Pamela  Ronald, 
University  of  California,  Davis,  CA, 
announced  the  public  availability  of 
a variety  of  libraries,  including  those 
on  Bacterial  Artificial  Chromosomes 
and  cosmids. 

Physical  Mapping 

Physical  mapping  was  again  high- 
lighted in  the  Arabidopsis  workshop. 
Caroline  Dean  (John  Innes  Institute, 
Norwich,  England)  and  Howard 
Goodman  (Massachusetts  General 


Hospital)  reported  that  chromo- 
somes 4 and  5 are  nearing  comple- 
tion in  their  joint  effort  to  integrate 
the  two  YAC  and  cosmid  maps.  A 
new  YAC  library  developed  by 
David  Bouchez  should  help  in 
developing  the  integrated  physical 
map.  Several  thousand  Arabidopsis 
cDNAs  have  been  sequenced  by  the 
French  EST  project.  Michel  Delseny, 
(CNRS,  Perpignan,  France)  who 
reported  on  the  project,  indicated 
that  the  cDNA  sequences  have  been 
deposited  in  the  public  database 

m 

"He  defined  this  era  as 
a new  voyage  of  the 
Beagle" 

m 

EMBL. 

Resources 

Plant  Genome  II  provided  partici- 
pants with  information  on  useful 
technologies  and  resources.  The 
latest  developments  in  the  plant 
genome  databases  were  outlined,  as 
well  as  computational  tools  for 
mapping  and  sequence  analysis. 
Database  demonstrations  with  a live 
Internet  link  were  available  through- 
out the  meeting,  allowing  hands-on 
experience  for  interested  researchers. 
Electronic  BIOSCI  newsgroups  were 
the  focus  of  several  workshops 
organized  by  Dave  Kristofferson 
(Intelligenetics,  Mountain  View,  CA). 

QTL  Experimental  Design 

Quantitative  trait  loci  (QTL)  analysis 
was  examined  with  attention  to 
experimental  design  and  analysis. 


Dave  Webb,  Pioneer  Hi-Bred, 
Johnston,  IA,  looked  at  soybean  cyst- 
nematode  resistance;  one  soybean 
introduction  was  found  to  have  more 
resistance  than  any  other  soybean 
tested  to  date.  Three  resistance  loci 
were  identified;  with  this  informa- 
tion, the  effect  of  population  size  in 
detecting  the  traits  was  tested.  Large 
sample  populations  were  found  to  be 
essential  in  finding  and  mapping 
these  traits.  The  minimum  sample 
population  size  is  200. 

The  need  for  large  sample 
populations  was  again  emphasized 
by  Karl  Lark,  University  of  Utah,  Salt 
Lake  City.  Lark  found  that  special- 
ized statistical  methods  and  graph- 
ing were  needed  to  identify  many 
important  loci.  Specifically,  Lark 
identified  in  interacting  traits  a 
condition  called  epistasis.  One  trait 
measured  on  its  own  had  no  effect  on 
plant  height.  This  same  trait  was 
found  to  interact  with  another  plant 
height  QTL  and  could  explain  25%  of 
the  plant  height  variation.  The  basis 
of  Lark's  technique  is  to  use  large 
population  sizes  and  to  conduct 
pairwise  comparisons  of  loci  in 
plants  with  extreme  phenotypes. 

The  results  are  graphed  and  epistatic 
interactions  are  then  identified. 

According  to  Thomas 
Cheesbrough,  South  Dakota  State 
University,  Brookings,  this  type  of 
analysis  will  be  essential  to  studying 
the  genes  of  such  metabolic  path- 
ways as  oil  production,  because  each 
enzyme  is  highly  interdependent  on 
the  gene  products  of  the  entire 
metabolic  chain. 

Mapping  Technologies 

Mapping  technologies  were  featured 
in  several  talks  and  posters  through- 
out the  conference.  Perry  Cregan, 
USD  A,  ARS,  Beltsville,  MD,  and 


14 


Probe 


Volume  4,  No.1/2 


Touching  Base  with  Lisa  Lorenzen 

Soybase  News 


Dr.  Lisa  Lorenzen 

Department  of  Zoology  and  Genetics 
Iowa  State  University 
Ames,  I A 

he  Soybase  staff  has 
been  concentrating  on 
the  metabolic  portion  of 
the  database.  Informa- 
tion (current  through  summer  1994) 
on  the  enzymes  and  reactions/ 
pathways  involved  in  nitrogen 
metabolism  have  been  entered  and 
became  available  August  15,  1994. 

The  complete  set  also  will  include 
references  for  all  cited  articles  and 
books. 

The  information  on  Fatty  Acids 
is  50  percent  collected  and  entered 
and  is  slated  for  completion  early  in 
1995.  A section  on  nodulins  has 
been  added. 


The  Pathology  section  continues 
to  grow,  with  downy  mildew, 
powdery  mildew,  frogeye  leaf  spot, 
potato  mosaic  virus,  cowpea  mosaic 
virus,  and  stem  canker  being  the 
latest  additions.  Soybase  staff 
members  have  collected  and  entered 
information  from  the  literature  on 
both  quantitative  and  qualitative 
trait  linkages  to  molecular  markers. 
We  wish  to  encourage  anyone  who 
has  data  that  he /she  wants  to 
include  in  Soybase  to  contact  us  at 
the  E-mail  address  listed  below.  We 
would  be  happy  to  include  your 
data. 


Conference— cont.  from  page  13 


others  reported  on  the  continued 
success  with  simple  sequence  repeats 
(SSR).  The  SSRs  are  small  sequence 
patterns  which  are  repeated  at 
variable  lengths.  The  variable  length 
of  the  repeats  provides  a means  to 
identify  varieties  and  individuals; 
tools  needed  by  crop  breeders  and 
geneticists.  In  addition  to  SSR 
technology,  amplified  fragment 
length  polymorphism  (AFLP),  a 
related  new  technology,  was  re- 
ported by  Drs.  Pieter  Vos  and  Marc 
Zabeau,  KeyGene,  Wageningen,  The 
Netherlands. 

AFLP  will  provide  markers  for 
those  map  regions  which  other 
markers  have  not  successfully 
bridged.  The  AFLP  technique  has  the 


capacity  to  exploit  multiple  forms  of 
variation  within  the  genome.  The 
new  technology  described  by  Vos  is 
still  a long  way  from  direct  applica- 
tion by  plant  breeders,  as  discussed 
at  the  International  Triticale  Map- 
ping Initiative  meeting  held  in  San 
Diego  in  conjunction  with  the  Plant 
Genome  II  conference. 

Plant  Genome  III 

Plant  Genome  III  will  be  held  Janu- 
ary 15-19,  1995,  in  San  Diego,  CA. 
Sessions  will  address  all  aspects  of 
mapping,  from  QTLs  to  the  latest 
molecular  marker  technologies, 
instrumentation,  and  gene  isolation. 
For  more  information  or  program 


Thinking  toward  an  integrated 
legume  database,  the  Soybase  staff 
has  assisted  in  the  initiation  of  four 
new  ACE-type  databases:  alfalfa, 
peanuts,  common  bean,  and  cool 
season  food  legumes  such  as  lentil 
and  chickpea.  These  databases  are 
being  administered  under  the 
supervision  of  Dr.  Daniel  Skinner, 
USDA-ARS,  Kansas  State  University; 
Dr.  Gary  Kochert,  University  of 
Georgia;  Dr.  Phil  McClean,  North 
Dakota  State  University;  and  Dr. 

Fred  Muehlbauer,  USDA-ARS, 
Washington  State  University;  respec- 
tively. 

For  additional  information  on 
Soybase,  or  for  instructions  on  how 
to  contact  one  of  the  new  legume 
databases,  please  send  inquiries  to 
curator@mendel.  agron.iastate.edu. 

♦ 


suggestions,  contact  Jerome  Miksche 
or  Stephen  Heller,  USDA/ARS, 
BARC-W,  Bldg.  005,  Room  331-C, 
Beltsville,  MD  20705  USA.  (See 
article,  "Announcing  Plant  Genome 
III"  in  this  issue.) 

♦ 


Probe 


July  1993  -July  1994 


The  Class  of  1993  Plant  Genome 
Grant  Recipients 


G.  Agrios,  P.  Chourey 
University  of  Florida 

Molecular  and  Physiological  Genetics  of  Sucrose 
Metabolizing  Enzymes  in  Maize  Endosperm 


M.  Alleman 
Duquesne  University 

Regulatory  Mutants  of  the  Maize  R Locus 


J.  Anderson,  F.  Francl 

North  Dakota  State  University  of 

Agriculture  & Applied  Science 

Molecular  Mapping  of 
Tan  Spot  Resistance 
Genes  in  Wheat 


Z.  Avramova 
Purdue  Research  Foundation 
Nuclear  Matrix  and  Matrix- 
Attachment  Regions  (MARs) 
in  Higher  Plants 


P.  Baenziger,  Y.  Yen 
Nebraska  Agricultural  Experiment  Station, 

University  of  Nebraska 

Exploring  the  Interface  of  Qualitative  and  Quantitative 
Genetics 


W.  Baird,  A.  Abbott,  R.  Ballard,  W.  Bridges 
Clemson  University 

Chromosome  Mapping  in  Peach  and  its  Application  to 
Fruit  Quality  Maintenance 


G.  Bates 

Florida  State  University 

Targeting  of  DNA  Integration  to  Specific  Sites  in  Plant 
Chromosomes 


J.  Birchler 

The  Curators  of  the  University  of  Missouri 
Molecular  Analysis  of  Maize  Centromeres 


D.  Bisaro 

Ohio  State  University  Research  Foundation 

Molecular  Mechanisms  of  Geminivirus  Replication 


T.  Blake,  L.  Talbert 
Montana  State  University 

Molecular  Markers  to  Correct  Germplasm  Deficiencies 
in  Wheat  and  Barley 


W.  Briggs 

Gordon  Research  Center 

1993  Gordon  Research  Conference 

on  Plant  Molecular  Biology 
Signal  Transduction  and 
Membrane  Proteins 

R.  Brown,  I. 
Saxena 

University  of  Texas 
Molecular  Analysis  of 
Cellulose  Biosynthesis  in 
Acetobacter  xylinum 


S.  Brown,  H.  Aldwinckle, 
N . Weeden 

Cornell  University 
Genome  Mapping  & Gene  Tagging  in  Apple 

M.  Bustos 

University  of  Maryland,  Baltimore  County 
Hormonal  Control  of  Gene  Expression  During  Seed 
Maturation 

R.  Cantrell 

Agricultural  Experiment  Station 

Board  of  Regents  of  New  Mexico  State  University 

Use  of  RAPD  Markers  to  Determine  the  Genetic 
Diversity  of  Gossypium  Germplasm  Derived  from 
Interspecific  Hybridization 

A.  Cheung 
Yale  University 

Molecular  and  Cellular  Analysis  of  a Pro-  and  Cys-rich 
Protein  Family  in  the  Pistil 


16 


Probe 


Volume  4,  No.1/2 


J.  Chory 

Salk  Institute  for  Biological  Studies 

Molecular  and  Genetic  Analysis  of  Arabidopsis  DET2 

Gene 

J.  Colbert 

Iowa  State  University  of  Science  and  Technology 
Phytochrome  mRNA  Degradation:  Cis-elements, 
Factors,  and  Pathway 

S.  Colby 

Individual  Awardee 

Regulation  of  Glandular  Trichome-based  Insect 
Resistance  in  Lycopersicon 

D.  Cook,  K.  VandenBosch 
Texas  A&M  Research  Foundation 

A Peroxidase  Gene  Induced  During  Nodule  Initiation 
in  Medicago  truncatula 

M.  Cordonnier-Pratt,  L.  Pratt 
University  of  Georgia  Research  Foundation,  Inc. 
Phytochrome  and  Potential  Photomorphogenic  Loci  in 
the  Grasses 

K.  Coschigano 

New  York  University 

Characterization  of  Two  Fd-GOGAT  Genes  and  Their 
Roles  in  Photorespiration 

P.  Cregan,  J.  Specht,  R.  Shoemaker 
USDA/ARS  Beltsville  Area 

An  Integrated  Microsatellite,  RFLP,  and  Conventional 
Linkage  Map  of  Soybean 

H.  Daniell 
Auburn  University 

Transformation  and  Foreign  Gene  Expression  Studies 
Using  the  Gene  Gun 

J.  Demski,  R.  Jarret 

University  of  Georgia  Research  Foundation,  Inc. 
Protoplast-Mediated  Transformation  of  Peanut  for 
Virus  Resistance 

R.  Dewey,  P.  Sisco,  D.  Danehower 
North  Carolina  State  University 

The  Glossy-15  Gene  of  Maize,  a Cell-Specific  Regula- 
tor of  Leaf  Epidermal  Traits 

J.  Doebley 

The  Regents  of  the  University  of  Minnesota 

Genetics  of  Inflorescence  Development  in  Maize  and 

Teosinte 


X.  Dong 

Duke  University 

Molecular  Genetic  Analysis  of  Systematic  Acquired 
Resistance  in  A.  thaliana 

H.  Dooner 

DNA  Plant  Technologies,  Inc. 

A Set  of  Maize  Lines  Carrying  Ac  at  Mapped  Locations 
Dispersed  in  the  Genome 

J.  Dvorak 

The  Regents  of  the  University  of  California 
Recombination  Between  Homoelogous  Chromosomes 
in  Wheat 

V.  Dzelzkalns 

Case  Western  Reserve  University 

Regulation  and  Mechanism  of  the  Self-Incompatibility 
Response  of  Flowering  Plants 

E.  Earle 

Cornell  University 

Chromosome-Specific  Libraries  of  Tomato 
S.  Farrand 

The  Board  of  Trustees  of  the  University  of  Illinois 

Cis-  and  Trans-Acting  Functions  Mediating  Ti  Plasmid 

Transfer 

R.  Ferl 

University  of  Florida 

Chromatin  Structure  and  Gene  Expression  in  Plant 
D.  Gallie 

The  Regents  of  the  University  of  California 
Isolation  of  RNA-Binding  Proteins  Involved  in  Regu- 
lating Translation 

S.  Gelvin 

Purdue  Research  Foundation 

Cell  Biology  of  T-DNA  Transfer  to  Plant  Cells 

P.  Gepts 

The  Regents  of  the  University  of  California 
Mapping  Genetic  Determinants  of  Host-Bacteria 
Interactions  in  Common  Bean 

R.  Gilbertson,  W.  Lucas 

The  Regents  of  the  University  of  California 

Molecular  and  Cellular  Analysis  of  Vascular  Function 

Using  a Phloem-Limited  Virus 

B.  Gill 

Kansas  State  University 

The  Sub-Ami  Aneuploids  of  Common  Wheat 


July  1993  -July  1994 


Probe 


17 


P.  Green 

Michigan  State  University 

Control  of  mRNA  Stability  in  Dicotyledonous  Plants 
A.  Grossman 

Carnegie  Institution  of  Washington 

The  Use  of  Cyanobacteria  to  Explore  Basic  Biological 

Processes  — 1993  Workshop 

L.  Hadwiger 

Washington  State  University 

Genetic  Engineering  of  Non-Host  Resistance  in  Plants 

L.  Hanley-Bowdoin 

North  Carolina  State  University 

DNA  Replication  in  Plants:  An  Accessory  Factor  for 

Geminivirus  Replication 

M.  Hanson 
Cornell  University 

Regulation  of  Synthesis  of  Plant  Mitochondrial 
Proteins 

D.  Harry,  D.  Neale 
PSWFRES,  USDA,  Forest  Service 

Codominant  PCR-based  Markers  for  Pines  and  Other 
Conifers 

G.  Hart 

Texas  A&M  Research  Foundation 
Construction  of  an  RFLP-Based  Genetic  Map  of 
Sorghum  Recombinant  Inbred  Lines 

T.  Hodges,  L.  Lyznik 
Purdue  Research  Foundation 
Gene  Targeting  of  Plant  Cells 

S.  Howell 

Boyce  Thompson  Institute  for  Plant  Research,  Inc. 
Isolation  of  Genes  Involved  in  Cytokinin  Responses 
in  Arabidopsis 

A.  Huang 

The  Regents  of  the  University  of  California 
Molecular  and  Cell  Biology  of  Oil  Bodies  in  Maize 
and  Brassica 

S.  Hulbert 

Kansas  State  University 

Analysis  of  the  Rpl  and  Rp3  Loci  of  Maize 

S.  Hulbert,  D.  Delaney,  B.  Gill 
Kansas  State  University 

Development  of  a High  Density  Chromosome  Map 
Using  Region-Specific  Libraries 


A.  Hung 

The  Regents  of  the  University  of  California 
Molecular  and  Cell  Biology  of  Oil  Bodies  in  Maize 
and  Brusque 

R.  Innes 

Indiana  University 

Molecular  Cloning  of  Disease  Resistance  Genes  from 
Arabidopsis  and  Soybean 

A.  Jagendorf 
Cornell  University 

Function  and  Regulation  of  Chloroplast  REC-A 
Protein 

M.  James,  A.  Myers 

Iowa  State  University  of  Science  and  Technology 

Isolation  and  Characterization  of  the  Maize  Gene 
Sugaryl,  a Determinant  of  Starch  Composition  in 
Kernels 

J.  Kermicle,  W.  Eggleston 

Board  of  Regents  of  the  University  of  Wisconsin  System 
Tests  for  Ac/Ds-Induced  Gene  Conversion  in  Maize 

R.  Kesseli 

University  of  Massachusetts 

Genome  Evolution  and  the  Organization  of  Disease 
Resistant  Genes  in  the  Coinpositae 

J.  Kikkert,  J.  Sanford 
Cornell  University 

Biological  Projectiles  for  Delivery  of  High  Molecular 
Weight  DNA  to  Plants 

A.  Kleinhofs 

Washington  State  University 

High  Resolution  Map  of  the  Barley  Sub-Telomeric 
Region  Including  Rpgl  Gene 

J.  Kohn 

The  Regents  of  the  University  of  California,  San  Diego 
QTL  Analysis  of  Developmental  Traits  in  Wild  Rice 

M.  Kuchka 
Lehigh  University 

Nuclear  Gene  Products  and  Chloroplast  Gene  Expres- 
sion 

C.  Lamb,  C.  Ryan 

Federation  of  American  Societies  for  Experimental 
Biology 

FASEB  Summer  Research  Conference:  Signal  Trans- 
duction in  Plants 


18 


Probe 


Volume  4,  No.1/2 


M.  Lee,  R.  Wise 

Iowa  State  University  of  Science  and  Technology 

Genetic  Organization  of  Resistance  to  Puccinia 
coronata  in  Avena 

B.  Liu,  D.  O'Malley,  F.  Bridgewater,  D.  Grattapaglia 
North  Carolina  State  University 

Genome  Map  Assisted  Plant  Breeding  (GMAPB)  for 
Forest  Tree 

S.  MacKenzie 

Purdue  Research  Foundation 

Arabidopsis  Mitochondrial  Genome  Alterations  in 
Response  to  Nuclear  Genotype 

P.  Maliga 

Rutgers,  The  State  University 

A Transgenic  Approach  to  Dissect  Light  Regulation  of 
the  Plastid  psbD/C  Operon 

P.  Maliga 

Gordon  Research  Center 

Gordon  Research  Conference  on  Plant  Cell  & Tissue 
Culture:  Plant  Transgenes-Tools  for  Discovery  and 
Design 

D.  McCarty,  I.  Vasil 
University  of  Florida 

Viviparous-1  Mediated  Repression  of  Alpha  Amylase 
Genes  in  Developing  Aleurone 

S.  McCouch 
Cornell  University 

High-Density  Genetic  Mapping  of  the  Rice  Genome 
Based  on  Sequence  Tagged  Microsatellite  Sites 

M.  Mutschler 
Cornell  University 

Genetic  Control  and  Field  Efficacy  of  Acylsugar 
Mediated  Multiple  Pest  Resistance 

J.  Nasrallah 
Cornell  University 

A Structural  and  Transcriptional  Analysis  of  the  S- 
Locus  Region  of  Brassica 

D.  Neale,  N.  Wheeler 
PSWFRES,  USDA,  Forest  Service 

Molecular  Marker  and  Quantitative  Trait  Mapping  in 
Douglas-Fir 

R.  Newton 

Texas  A&M  Research  Foundation 

Conifer  Transformation  with  Shoot  Apices  and 

Agrobacterium 


H.  Nguyen,  D.  Rosenow 
Texas  Tech  University 

Tagging  Drought  Tolerance  Traits  in  Grain  Sorghum 
Using  Molecular  Markers 

C.  Opperman,  M.  Conkling 
North  Carolina  State  University 
Characterization  of  a Nematode-Responsive  Plant 
Gene  Promoter 

P.  Ozias-Akins,  W.  Hanna 

University  of  Georgia  Research  Foundation,  Inc. 
Development,  Genomic  Diversity,  and  Gene  Expres- 
sion in  Aposporous  Genotypes 

G.  Phillips,  G.  Kuehn,  S.  Bagga 

Agricultural  Experiment  Station 

Board  of  Regents  of  New  Mexico  State  University 

Isolation  of  Genes  Coding  for  Plant  Polyamine  Bio- 
synthetic Enzymes 

G.  Powell,  A.  Abbott 

Clemson  University 

Characterization  of  the  12-Desaturase 

L.  Pratt,  M.  Cordonnier-Pratt 

University  of  Georgia  Research  Foundation,  Inc. 

Phytochrome  Gene  Family  in  Tomato 

C.  Qualset 

The  Regents  of  the  University  of  California 

Research  Collaboration  Group  on  Molecular  Mapping 

in  Wheat  and  Its  Relatives 

R.  Rajasekharan,  J.  Kemp 

Agricultural  Experiment  Station 

Board  of  Regents  of  New  Mexico  State  University 

Lysophosphatidic  Acid  Acyltransferase:  Enzyme  and 

Gene  Isolation  from  Soybean 

L.  Ream 

Oregon  State  University 

A Multifunctional  DNA  Binding  Protein  Required  for 
Gene  Transfer  to  Plants 

K.  Redman,  M.  Johnson 
University  of  Alabama 

A Novel  Mechanism  for  Ribosomal  Protein  Modula- 
tion: On/Off  Splicing 

P.  Ronald 

The  Regents  of  the  University  of  California 
Map-based  Cloning  in  Rice 

L.  Rowland 

USDA/ARS  Beltsville  Area 


July  1993  -July  1994 


Probe 


19 


Tagging  Genes  which  Control  Chilling  Requirement 
in  a Woody  Perennial 

M.  Saghai  Maroof 

Virginia  Polytechnic  Institute  and  State  University 
Assessment  of  Barley  Germplasm  Using  Nuclear  and 
Organellar  Molecular  Markers 

R.  Schmidt,  M.  Yanofsky 

The  Regents  of  the  University  of  California,  San  Diego 
An  Analysis  of  Floral  Regulatory  Genes  in  Maize 

S.  Scofield 

The  Regents  of  the  University  of  California 
Promoters  to  Express  Ac  Transposase  for  Efficient 
Tagging  Systems 

E.  Signer 

Massachusetts  Institute  of  Technology 
Repeat-Induced  Gene  Silencing  in  Arabidopsis 

S.  Strauss,  W.  Rottmann 
Oregon  State  University 

Floral  Homeotic  Genes  for  Genetic  Engineering  of 
Sterility  in  Populus 

C.  Stuber 

USDA/ARS  South  Atlantic  Area 

Stability  of  QTL  Mapping  in  Maize  Under  Varying 

Environmental  Stresses 

T.  Sullivan 

Board  of  Regents  of  the  University  of  Wisconsin  System 
Molecular  and  Biochemical  Analysis  of  the  Maize 
Brittle-1  Gene 

S.  Sun 

University  of  Hawaii,  Manoa 

Genetic  Transformation  of  Grain  Legumes  for  Im- 
proved Protein  Quality 

R.  Thornburg 

Iowa  State  University  of  Science  and  Technology 

Selection  for  Second  Site  Mutations  in  the  Wound- 
Induction  Pathway 

J.  Trumble 

The  Regents  of  the  University  of  California 
Transgenic  Insect-Resistant  Brassica  with  Glossy  Wax 
Genes  from  Arabidopsis 

R.  Vierstra 

Board  of  Regents  of  the  University  of  Wisconsin  System 
Molecular  and  Biochemical  Analysis  of  Ubiquitin 
Conjugating  Enzymes  in  Higher  Plants 


C.  Weil 

University  of  Idaho 

Transposable  Element-Mediated  Dissection  of  Protein 
Structure  and  Function 

R.  Wing,  A.  Paterson 
Texas  A&M  Research  Foundation 
Physical  Mapping  and  Map-Based  Cloning  in  Polyp- 
loids: Cotton  as  a Model  System 

R.  Wise 

USDA/ARS  Mid-West  Area 

High  Resolution  Mapping  of  the  Ml-a  Disease  Resis- 
tance Locus  in  Barley 

S.  Yang 

The  Regents  of  the  University  of  California 

ACC  Malonyltransferase:  Isolation,  Characterization 

and  Molecular  Cloning 

J.  Yoder 

The  Regents  of  the  University  of  California 
Transposon  Mutagenesis  in  Tomato 

R.  Zielinski 

The  Board  of  Trustees  of  the  University  of  Illinois 
Molecular  Characterization  of  CaBP-22,  a Leaf-Spe- 
cific, EF-Hand  Ca2+  -Binding  Protein 


IMPORTANT 


1995  NKI 
Grant 

Deadlines 

Announced 

See  page  8 of  this  issue 


Probe 


Volume  4,  No.1/2 


Touching  Base  with  Bradley  Sherman 


A Primer  on  Images  and  the  Internet 


Bradley  K.  Sherman 

Institute  of  Forest  Genetics 

USDA  Forest  Service 

P.O.  Box  245  Berkeley , CA  94701 

bks@s27w007.pswfs.gov 


High-speed  digital  networks  and 
computers  with  graphical  capabilities 
have  made  it  possible  to  retrieve  and 
view  images  almost  as  easily  as  one 
views  purely  textual  material.  This 
article  is  intended  to  be  a quick 
introduction  to  this  subject  with 
some  pointers  to  other  resources. 

What  Is  a Digital  Image? 

Digitally  stored  images  are  analo- 
gous to  a paint-by-number  kit.  The 
image  is  composed  of  rectangular 
regions  called  pixels,  and  each  pixel 
is  assigned  a number.  Each  number 
corresponds  to  a color  or  shade  of 
grey.  In  order  to  reconstruct  an 
image  for  viewing,  one  needs  both 
the  values  for  each  pixel  and  a 
legend  which  maps  the  numerical 
values  to  a color.  Image  files  on 
computers  typically  contain  both  the 
array  of  pixel  values  and  the  color 
map. 

Digital  images  usually  contain 
less  information  than  a correspond- 
ing image  captured  with  conven- 
tional photographic  methods. 
Photographic  film  is  an  excellent 
repository  for  information.  One 


35mm  slide  can  easily  hold  one 
100  million  bytes  of  data.  This 
makes  the  digital  storage  of 
images  somewhat  problematic. 
Even  if  the  cost  of  storage  were 
not  a factor,  the  time  involved  in 


"What  is  the  use  of  a 
book , " thought  Alice , 

"without  pictures  or 
conversations? " — 
Lewis  Carroll 


retrieving  and  displaying  very 
large  stored  images  would  be 
prohibitive.  All  common  conver- 
sion of  photographic  data  to 
digital  data  involves  the  loss  of 
some  information.  Once  the 
images  are  digitized,  they  are 
often  compressed.  Some  compres- 
sion methods  result  in  further 


information  loss.  There  is  always  a 
tradeoff  between  faithfulness  of 
reproduction  and  amount  of  storage 
space  required.  Even  if  storage  were 
free,  it  would  be  advantageous  to 
keep  the  image  size  small  for  rapid 
transferal  and  viewing.  This  dynamic 
has  led  to  many  different  formats  for 
computer  images,  each  with  advan- 
tages and  disadvantages.  The  deci- 
sion about  which  format  to  use  is 
highly  dependent  on  the  application, 
and  on  the  hardware  that  will  be 
used.  On  the  Internet,  exchanged 
images  tend  to  follow  two  main 
formats:  GIF  and  TIFF.  These  for- 
mats, particularly  the  latter,  have 
many  variations.  JPEG  images  are  a 
ubiquitous  TIFF  variant  (JFIF). 

In  addition  to  these,  different 
computer  types  can  have  their  own 
internal  formats,  customized  for  their 
particular  hardware.  The  PICT 
format  on  Macintosh  is  an  example 
of  this. 

Software 

There  is  software  available  for  Unix, 
Macintosh  and  Intel-based  platforms 
that  will  allow  you  to  view  down- 
loaded images.  You  may  retrieve 
them  using  anonymous  ftp  and 
experiment  with  them  for  free.  Some 
of  the  software  is  shareware,  and  the 
author  expects  some  small  compen- 
sation if  you  like  the  software  and 
continue  to  use  it.  Sources  for  image 


July  1993  -July  1994 


Probe 


21 


viewing  software  for  major  platforms 
are  listed  at  end  of  this  article. 

Hardware 

Workstations  such  as  those  from  Sun 
Microsystems  or  Silicon  Graphics 
were  developed  with  advanced 
graphical  uses  in  mind.  High-end 
Macintoshes  and  PC-clones  are 
adequate,  however.  Monitor  screens 
should  be  at  least  15  inches,  and 
graphics  hardware  may  be  helpful  or 
necessary.  Image  analysis  can  be 
slow  even  on  very  fast  computers. 

Typical  Image  Sizes  and 
Network  Bandwidth 

Images  seen  on  the  Internet  can 
range  in  size  from  hundreds  to 
millions  of  bytes.  A four-megabyte 
file  is  not  uncommon.  Internet 
connections  can  be  characterized  by 
the  bandwidth  of  the  connection. 
Typical  bandwidths  are  9.6,  14.4,  and 
56  kilobaud.  Major  nodes  may  have 
one  megabaud  connections  or  better. 
Bytes  per  second  can  be  approxi- 
mated by  dividing  the  baud  rate  by 
10.  A 56  kilobaud  channel  will 
transfer  about  5,600  bytes  per 
second. 

A typical  100  kilobyte  image  will 
take  at  least  1 second  to  move  across 
a 1 megabaud  channel  and  more  than 
17  seconds  across  1 of  56  kilobaud.  In 
addition,  the  presentation  of  the 
image  on  the  computer  screen  may 
take  seconds  once  the  image  has  been 
transferred. 

To  the  user,  large  images  or 
slow  network  connections  will  be 
seen  as  delays.  An  occasional  10- 
second  delay  after  pressing  a key  or 
mouse  can  be  tedious  in  an  interac- 
tive environment,  arguing  for  very 
high  speed  network  connections  or 


using  smaller  image  files.  Compres- 
sion techniques  can  be  very  useful, 
particularly  when  animated  images 
are  transferred. 

Image  Capture 

There  are  at  least  two  sorts  of  devices 
which  one  can  use  to  digitize  an 
image.  Both  make  use  of  charge- 
coupled  devices  (CCD)  which  use 
quantum  effects  to  transduce  light  to 
a pattern  of  electrical  signals.  Scan- 
ners have  a linear  array  of  CCDs 
which  are  mechanically  moved 
across  a flat  image  (in  the  manner  of 
a xerographic  machine).  CCD 
cameras  have  a two-dimensional 
array  of  devices  so  that  the  entire 
image  is  captured  at  once.  CCD 
cameras  use  conventional  optics  and 
hence  can  be  used  to  take  pictures  of 
3-D  objects. 

CCD  devices  are  susceptible  to 
thermal  noise;  they  will  produce 
small  random  signals  even  in  abso- 
lute darkness.  To  increase  signal-to- 
noise  ratios,  the  devices  can  be 
cooled.  Some  CCD  cameras  come 
with  refrigeration  units  for  this 
purpose.  CCD  cameras  are  more 
expensive,  but  allow  for  a higher 
throughput  in  a production  setting. 

Commercial  photographic 
processing  labs  commonly  have 
equipment  to  directly  digitize  color 
slides.  Copy  stores  will  often  have 
scanners  available  for  rental.  These 
work  quite  well  with  color  or  black- 
and-white  prints. 

Where  To  Get  Image  Viewing 
Software 

All  of  this  software  may  be  retrieved 
using  anonymous  ftp. 


Macintosh 

JPEGView 

ftp  to  sumex.aim.stanford.edu 
directory:  /info-mac/app/ 

NIH  Image 

ftp  to  zippy.nimh.nih.gov 
directory:  /pub/nih-image/ 

Unix/ XI 1 
xv 

ftp  to  bongo.cc.utexas.edu 
directory:  /gifstuff/xwindows/ 
xloadimage 

ftp  to  bongo.cc.utexas.edu 
directory:  /gifstuff/xwindows/ 

Intel/ Windows3 
Lview 

ftp  to  oak.oakland.edu 
directory:  /pub/msdow/windows3/ 

For  More  Information 

These  Usenet  electronic  conferences 
are  sources  of  useful  discussion: 
alt. binaries. pictures. misc 
comp. graphics 
alt. graphics. pixutils 

Lists  of  Frequently  Asked 
Questions  (FAQ),  with  answers,  for 
these  conferences  are  available  by 
anonymous  ftp  from  rtfnr.mit.edu. 


Probe 


Volume  4,  No.1/2 


Touching  Base  with  Mary  Polacco 


/VS&n.' 


Maize  Genome  Database,  a LJSDA- 
ARS  Plant  Genome  Database 


Mary  Polacco,  Database  Developer 
Curtis  Hall,  University  of  Missouri 
Columbia,  MO  65211 

The  Maize  Genome  Database,  or 
MaizeDB,  is  curated  as  a Sybase 
database  at  the  University  of  Mis- 
souri-Columbia  and  provides  user- 
friendly,  Internet  access  to  the  maize 
genome  and  the  biology  of  maize. 

Information  includes  142 
genetic  maps,  with  4,864  mapped 
loci,  recombination  and  map  score 
data  (2,164  entries),  986  probes,  1,701 
genetic /cytogenetic  stocks,  7994 
locus  variations,  4,662  stock  pedi- 
grees, 5,600  bibliographic  references 
indexed  to  genetic  objects,  and 
addresses  of  maize  researchers. 

Gene  functionality  may  be 
queried  by  mutant  phenotype,  trait, 
confirmed  or  putative  gene  products, 
metabolic  pathways,  induction 
conditions  and  text  descriptions  of 
genes.  Work  is  in  progress  to  docu- 
ment quantitative  trait  loci.  Users 
with  Internet  connections  may  access 
the  data  by  several  procedures  that 
are  described  more  fully  below: 
guest  login,  gopher.  World  Wide 
Web,  or  file  transfer  of  a special 
graphics,  ACEDB  format. 

Data  is  input  from  various 
sources,  including  specially  format- 
ted, electronic  lab  notebooks  of 
researchers  who  focus  on  mapping 
or  mutant  characterization.  The 
contributors  are  international  and 


from  academic,  government,  and 
industrial  research  groups.  Informa- 
tion, especially  regarding  gene 
function  and  expression,  is  also  taken 
from  the  scientific  literature,  both 
electronic  and  printed. 

Connecting  To  Other 
Databases 

Using  Mosaic,  a type  of  free  software 
that  connects  users  to  information  on 
the  World  Wide  Web  (WWW),  users 
connected  to  MaizeDB  may  retrieve 
information,  including  images  and 
graphics,  from  other  databases 
around  the  world,  as  easily  as  if  the 
data  existed  in  the  MaizeDB  records. 
If  desired,  they  may  store  the  infor- 
mation on  their  machines. 

For  example,  while  browsing 
the  MaizeDB,  you  may  read  that  the 
function  of  maize  gene,  dpsl,  was 
confirmed  by  transgenic  complemen- 
tation of  E.  coli  mutations  in  dapl. 

By  clicking  on  dapl,  you  would 
retrieve  the  record  from  the  E.  coli 
Stock  Center  at  Yale  University. 
Clicking  on  the  EC  number  for  gene 
products  that  are  enzymes  connects 
you  to  the  ENZYME  database. 
ENZYME  describes  the  reaction,  and 
in  turn,  connects  to  all  Swiss-Prot 
entries  corresponding  to  that  EC 
number,  as  well  as  to  OMIM  (On-line 


Mendelian  Inheritance  in  Man). 

WWW  connectivity  requires 
precise  matching  of  records  in 
MaizeDB  to  records  in  other  data- 
bases around  the  world.  It  permits 
curators  of  distinct  datasets  to 
combine  data  in  a seamless  fashion 
without  actually  importing  the  data. 
The  abihty  to  extract  distinct  formats 
of  data  from  MaizeDB  makes  prepar- 
ing files  of  matching  identifiers 
relatively  easy,  so  that  external 
databases  may  use  the  WWW  to 
connect  to  us,  as  occurred  in  June 
1994  with  SwissProt. 

WWW/ MaizeDB  currently 
accesses  external  data  in  the  follow- 
ing databases: 

GenBank nucleotide  sequences 

SwissProt protein  sequences; 

connects  to  Prosite  (motifs,  signature 
sequences),  MedLine,  EMBL 

dbEST random  cDNAs  partial 

sequences,  with  periodically 
updated  homology  searches 
E.  coli  Stock 

Center E.  coli  genetic  stocks,  map 

ENZYME reactions,  comments; 

connects  to  SwissProt,  OMIM 

AAtDB Arabidopsis  Genome 

Database 

Accessing  MaizeDB- 
Requirements 

Guest  login  only  requires  that  your 
machine  have  Internet  connectivity, 
direct  or  indirect.  Modem  connec- 
tions are  supported,  as  are  connec- 
tions using  any  computer,  including 


July  1993 -July  1994 


Probe 


23 


PC,  Macintosh,  and  Unix, 
guest  login  protocol: 
telnet  teosinte.agron.missouri.edu 
login:  guest 
password:  corncob 
Guest  login  provides  access  to: 

• gopher 

• MaizeDB,  a Sybase  database; 
access  provided  to  users  with 
either  X-Window  or  vtlOO 
emulation 

• Lynx,  a WWW  browser  that 
does  not  require  an  X-Window; 
it  does  not  support  mouse- 
capability 

• help 

Guests  are  encouraged  to  leave 
comments  on  the  Note  form  of  the 
database.  While  not  required,  leaving 
your  e-mail  address  will  permit  us  to 
contact  you  directly  for  further 
clarification. 

NOTE:  Users  with  X-Windows 
(this  is  not  the  same  as  Microsoft 
Windows)  software  will  enjoy  the 
most  user-friendly  access  to  the 
database.  If  connecting  by  modem,  the 
X-Window  will  not  function,  and 
users  should  select  the  vtlOO  emula- 
tion. 

NOTE:  If  using  the  vtlOO  emula- 
tion of  MaizeDB /Sybase,  type  "r" 
while  holding  down  the  "control"  (aka 
"CTRL")  key  to  access  the  commands 
required  to  query  or  browse  the 
database.  The  command  utilities  are 
described  in  more  detail  in  the  "help" 
option  that  appears  or  after  successful 
login  as  a guest. 

Gopher 

Gopher  makes  available  hierarchical 
collections  of  information  across  the 
Internet.  Gopher  client  (user)  soft- 
ware provides  easy  access  to  all 
gopher  data  servers.  All  words  in  a 
record,  except  commonly  used 
words,  are  indexed  and  thus  may  be 


used  to  query  records. 

Free  gopher  client  software  for 
Unix,  PC,  or  Macintosh  machines  is 
available  by  anonymous  ftp  (file 
transfer  protocol)'  from 
boombox.umn.edu.  Once  installed, 
open  server 

teosinte.agron.missouri.edu,  port  70 
or  use  gopher  to  find  us  by  location 
in  Columbia,  Missouri.  On-line  help 
is  provided  by  the  gopher  software 
and  is  in  a file  on  the  MaizeDB 
gopher  server. 

World  Wide  Web  (WWW) 

WWW  is  a hypermedia  retrieval 
system  which  allows  users  to 
traverse  on-line  documents  by 
clicking  on  hyperlinks-terms,  icons, 
or  images  that  point  to  other  related 
documents.  Hyperlinks  permit 
retrieval  of  any  document  anywhere 
on  the  Internet.  Retrieved  "docu- 
ments" may  include  text  files, 
graphics,  and  videos.  Connecting  to 
the  WWW  currently  works  best  if 
users  have  access  to  Mosaic  software 
installed  on  a Unix  machine. 
Macintosh  and  PC  Mosaic  software 
are  rapidly  approaching  the  capabil- 
ity of  the  Unix  version. 

Users  without  WWW  software 
may  access  the  WWW-linked  format 
of  MaizeDB  by  selecting  the  Lynx 
option  after  "guest  login." 

Mosaic  software  supports 
mouse  capability  and  is  available 
without  charge  by  anonymous  ftp 
from  ftp.ncsa.uiuc.edu.  The  Unix 
version,  but  not  the  Macintosh  or  PC 
version,  requires  an  X-Window  on 
the  user's  machine;  it  will  require  a 
systems  administrator  to  install.  To 
access  MaizeDB  from  Mosaic  soft- 
ware, use  our  WWW  address, 
otherwise  known  as  URL  or  uniform 
resource  locator: 

http:  / / teosinte.agron.missouri.edu  / 
top.html 


The  WWW  formatted  data  is 
dynamically  extracted  from  the  most 
current  version  of  the  database, 
which  is  continuously  updated. 

ACEDB  Format 

This  is  a special  graphical  format  and 
requires  a UNIX  machine.  The 
database  may  be  retrieved  by  anony- 
mous ftp  from  the  National  Agricul- 
tural Library,  probe.nalusda.gov  in 
directory  pub/ maize.  This  format  is 
static,  and  periodically  extracted 
from  MaizeDB.  It  does  not  support 
the  robust  queries  of  the  Sybase 
database,  accessible  by  the  guest 
login  service. 

ANONYMOUS  FTP  requires 
that  the  user  have  ftp  or  file  transfer 
software  to  connect  to  another 
machine.  Once  connected,  login  as 
"anonymous"  and  use  your  e-mail 
address  as  the  password.  If  using  a 
Unix  machine,  type:  cd  pub  /maize, 
and  to  transfer  the  database,  type: 
get  mace. tar. Z 

History  Of  MaizeDB  Design: 
Some  Landmarks 

Fall  1991 

First  prototype  MaizeDB  operational. 
Some  24,000  records  created  the  first 
6 months,  largely  from  data  summa- 
ries in  the  Maize  Genetics  Coopera- 
tion Newsletter  (MNL),  volume  65. 
The  database  currently  contains  over 
78,000  records. 

December  1992 

First  public  access  to  the  data,  a 
gopher  server  established.  First 
access  was  100-200  connections/ 
month,  and  has  grown  to  over  1,000 
connections/ month. 

March  1993 

Maize  Gene  List,  MNL,  vol  67,  pp. 
134-15,  extracted  from  MaizeDB 
Version  2 of  MaizeDB  implemented. 


Probe 


June  1993 

ACEDB  formatted  data  extracted 
from  MaizeDB. 

August  1993 

Tool  developed  for  loading  refer- 
ences from  PC  and  Macintosh 
reference  manager  formats. 

December  31,  1993 

Guest  login  to  MaizeDB  established. 

Winter  1994 

MaizeDB  placed  on  the  World  Wide 
Web;  currently  there  are  600-1,100 
connections/ week. 

WWW  connections  made  to  external 
databases,  listed  above. 


March  1994 

Genetic  indexing  of  1993  references 
extracted  from  the  MaizeDB,  pub- 
lished as  hardcopy  in  MNL,  vol  69. 
pp  148-153.  Information  was  indexed 
to  chromosome,  gene  or  allele  and 
trait. 

June  1994 

SwissProt  connects  to  MaizeDB 
using  a file  extracted  from  MaizeDB 
per  specifications  of  SwissProt 
curators. 


Volume  4,  No.1/2 


Developers  and  Curators  of 
the  Database  Include: 

E.H.  Coe  (PI),  P.  Byrne,  G.  Davis,  D. 
Hancock,  M.  Polacco  (Columbia,  MO) 
M.  Berlyn,  S.  Letovsky  (New  Haven, 
CT) 

C.  Fauron  (Salt  Lake  City,  UT) 

S.  Rodermel,  C.  Wetzel  (Ames,  IA) 

M.  Sachs  (Urbana,  IL) 

For  further  help  in  accessing  the 
database,  please  e-mail 
db_requeskafeosinte.agron.missouri.edu 
or  contact  Denis  Hancock, 

(314)  882-1722  (phone) 

(314)  874-4063  (FAX)  + 


Introducing  Dr.  Edward  Kaleikau 


Dr.  Edward  K. 

Kaleikau  has  as- 
sumed responsibility 
as  program  director 
for  the  Plant  Genome  Program  of 
the  USDA  National  Research 
Initiative  Competitive  Grants 
Program  (NRICGP).  In  this  position. 
Dr.  Kaleikau  coordinates  the  com- 
petitive grant  review  process  for  the 
Plant  Genome  Program.  His  respon- 
sibilities include  selecting  and 
working  with  members  of  the 
review  panel  in  conjunction  with  the 
panel  manager,  as  well  as  handling 
other  review  assignments  as 
needed. 

In  addition.  Dr.  Kaleikau  is  co- 
chairman  of  the  Plant  Genome 
Steering  Committee  along  with  Dr. 


Jerome  Miksche,  and  serves  on  the 
USDA  Biotechnology  Research 
Subcommittee. 

Dr.  Kaleikau,  a native  of 
Hawaii,  received  his  B.S.  degree  in 
biology /chemistry  (1981)  from 
Graceland  College  in  Lamoni,  IA  and 
PhD.  degree  in  plant  genetics  (1988) 
from  Kansas  State  University.  His 
Ph.D.  dissertation  investigated  the 
inheritance  and  chromosomal 
mapping  of  genes  controlling  in  vitro 
tissue  culture  response  in  wheat.  Dr. 
Kaleikau  developed  his  interest  in 
plant  genetics  during  an  internship  at 
the  Arco  Plant  Research  Institute  in 
Dublin,  CA,  after  graduation  from 
college.  He  gathered  further  experi- 
ence as  a technician  for  Advanced 
Genetic  Sciences  in  Manhattan,  KS. 


Prior  to  joining  the  NRICGP, 
Dr.  Kaleikau  received  postdoctoral 
training  at  Stanford  University, 
where  he  was  awarded  fellowships 
from  both  the  National  Institute  of 
Health  and  National  Science 
Foundation  to  study  the  regulation 
of  transcription  initiation  and 
termination  of  rice  mitochondrial 
genes. 

Dr.  Kaleikau  can  be  contacted 
as  follows: 

Internet: 

EKLEIKAU@DARTH.ESUSDA.GOV 
Phone:  (202)  401-5114 
USDA /CSRS/ NRICGP 
901  D Street,  SW 
Aerospace  Bldg,  Rrn  323 
Washington,  DC  20250-2241  ♦ 


July  1993  -July  1994 


Probe 


25 


Announcing  Plant  Genome  III 

Meeting 


Building  on  the  successes  of  Plant  Genome  I and  II,  we 
are  pleased  to  announce  that  the  Plant  Genome  III 
meeting  will  be  held  on  January  15-19,  1995,  in  San 
Diego,  CA. 

Session  Topics: 

1.  Comparative  Genetic  Mapping 

2.  Isolation  and  Transformation  of  Agriculturally 
Important  Genes 

3.  Instrumentation/Technology 

4.  Applications  of  cDNA  Research 

5.  Chromosome  Structure 

6.  ALFPs/QTLs/Metabolic  Pathways 

In  addition  to  the  formal  sessions  and  posters 
during  the  week,  Sunday  afternoon  will  feature  a com- 
puter workshop  on  Genome  Information  Tools  and 
Resources.  The  workshop  will  provide  a view  of  some  of 
the  existing  software  used  to  maintain  and  analyze 
genomic  information.  The  talks  at  this  workshop  are 
designed  to  give  the  average  plant  molecular  biologist  an 
idea  of  available  resources  and  their  capabilities.  "Hands 
on"  computer  sessions  will  also  be  available  throughout 
the  week.  Don't  miss  this  opportunity  for  individualized 
training. 

Additional  Workshops: 

Sunday,  January  15:  9:00  am  - 6:00  pm 
International  Consortium  for  Sugar  Cane  Biotechnology 
Organized  by  James  Irvine  (JIRVINE@TAMU.EDU) 

Tuesday,  January  17:  3:30  pm  - 6:00  pm 

1.  Pine  Tree  - Part  I 
Organized  by  Dave  Neale 
PBN@S27W007.PSWFS.GOV) 

2.  Rice 

Organized  by  Susan  McCouch 


(SUSAN_MCCOUCH@QMRELAYMAIL.CORNELL.EDU) 

3.  BIOSCI 

Organized  by  Dave  Kristofferson 
(KRISTOFF@NET.BIO.NET) 

4.  Arabidopsis 

Organized  by  Caroline  Dean 
(ARABIDOPSIS@BBSRC.AC.UK) 

Howard  Goodman 

(GOODMAN@>FRODO.MGH.HARV  ARD.EDU) 

5.  Barley 

Organized  by  Patrick  Hayes 
(HAYESP@CSS.ORST.EDU) 

Tuesday,  January  17:  7:30  pm  - 10:00  pm 

1.  Pine  Tree  - Part  II 
Organized  by  Dave  Neale 
PBN@S27W007.PSWFS.GOV) 

2.  Grass  Genome  Integration 

Organized  by  Jeff  Bennetzen 
(MAIZE@BILBO.BIO.PURDUE.EDU) 

Michael  Gale  (JEFFERY@BBSRC.AC.UK) 

3.  Nomenclature 
Organized  by  Carl  Price 
(PRICE@MBCL.RUTGERS.EDU) 

Ellen  Reardon 

(REARDON@CCIT.ARIZONA.EDU) 

4.  Tree  Fruit 

Organized  by  Sriyani  Rajapakse 
(SRIYANI_RAJAPAKSE@QUICKMAIL.CLEMSON.EDU) 

5.  BIOSCI  (Repeat  of  afternoon  session) 

Organized  by  Dave  Kristofferson 
(KRISTOFF@NET.BIO.NET) 

Wednesday,  January  18:  3:30  pm  - 6:00  pm 
1.  Maize 

Organized  by  Ed  Coe 

(ED@TEOSINTE.AGRON.MISSOURI.EDU) 


26 


Probe 


Volume  4,  No.1/2 


2.  Legumes 

Organized  by  Randy  Shoemaker 
(RCSSHOE@IASTATE.EDU) 

3.  ITMI 

Organized  by  Calvin  Qualset 
(ITMI@UCDAVIS.EDU) 

Olin  Anderson  (OANDERSON@PW.USDA.GOV) 
Pat  McGuire  (ITMI@UCDAVIS.EDU) 

Michael  Gale  (JEFFERY@BBSRC.AC.UK) 

4.  Tagging  Genes  for  Abiotic  Stress 
Organized  by  Henry  Nguyen  (806-742-1622) 

5.  Cotton 

Organized  by  Andrew  Paterson 
(AHP2343@BIOCH.TAMU.EDU) 

Abstract  Deadline: 

Abstracts  submissions  are  due  by  November  1,  1994.  All 
submitted  or  invited  poster  talks  will  be  one-page  long, 
using  forms  provided  by  the  PG-III  conference  organizer, 
Scherago  International.  The  PG-III  abstracts  will  be 
available  online  prior  (and  after)  to  the  meeting  at 
probe.nalusda.gov  via  gopher  and  the  WWW. 

Student  Travel  Grants: 

The  International  Society  for  Plant  Molecular  Biology  is 
again  sponsoring  four  student  travel  grant  awards.  For 
details  please  contact  Dr.  Stephen  Heller  by  E-mail. 

Location:  Town  & Country  Hotel 

500  Hotel  Circle  North 
San  Diego,  CA  92108 
Phone:  (619)  291-7131 
FAX:  (619)  291-3584 

Cost:  $300  advance  registration  up  to  December  1, 1994 
$350  after  December  1,  19.94  and  on-site 
$100  Student  (Pre-Ph.D)  registration 
(Requires  a letter  of  certification  from 
department  chairperson) 

All  registrations  include  one  copy  of  the  printed  confer- 
ence abstracts,  Monday-Thursday  continental  breakfasts, 
Sunday  evening  opening  reception,  Monday  evening 
wine  & cheese  reception,  and  Wednesday  evening 
dinner. 


PG-III  Co-Chairpersons: 

Stephen  Heller,  USDA/ARS,  Beltsville,  MD,  USA 
(SRHELLER@ASRR.ARS.USDA.GOV) 

Jerome  Miksche,  USDA/ARS,  Beltsville,  MD,  USA 
(JMIKSCHE@ASRR.ARSUSDA.GOV) 

Michael  Gale,  John  Innes  Centre,  Norwich,  UK 
(JEFFERY@BBSRC.AC.UK) 

Susan  McCouch,  IRRI,  Philippines 
(SUSAN_MCCOUCH@QMRELAY.MAIL.CORNELL.EDU) 

Conference  Co-sponsors: 

USDA,  Agricultural  Research  Service 
USDA,  National  Agricultural  Library 
Rockefeller  Foundation 

International  Society  for  Plant  Molecular  Biology 
John  Innes  Centre 

To  Register  Contact: 

Darrin  Scherago 
Scherago  International,  Inc. 

11  Penn  Plaza,  Suite  1003 
New  York,  NY  10001 
Phone:  (212)  643-1750 
FAX:  (212)  643-1758 

E-mail:  SCHERAGO@BIOTECHNET.COM  ♦ 

The  United  States  Department  of  Agriculture 
(USDA)  prohibits  discrimination  in  its  pro- 
grams on  the  basis  of  race,  color,  national 
origin,  sex,  religion,  age,  disability,  political 
beliefs,  and  marital  or  familial  status.  (Not  all 
prohibited  bases  apply  to  all  programs.) 

Persons  with  disabilities  who  require  alterna- 
tive means  for  communication  of  program 
information  (braille,  large  print,  audiotape, 
etc.)  should  contact  the  USDA  Office  of  Com- 
munications at  (202)  720-5881  (voice)  or  (202) 
720-7808  (TDD). 

To  file  a complaint,  write  the  Secretary  of 
Agriculture,  U.S.  Department  of  Agriculture, 
Washington,  D.C.  20250,  or  call  (202)  720-7327 
(voice)  or  (202)  720-1127  (TDD).  USDA  is  an 
equal  employment  opportunity  employer. 


Probe 


July  1993  -July  1994 


Calendar  of  Upcoming  Genome  Events 


Meetings 


September  18-22,  1994:  16th  International  Congress  on 
Biochemistry  and  Molecular  Biology,  New  Delhi, 
India.  Contact:  K.  Kooraram,  Magnet  World  Travel,  18- 
30  Clerkenwell  Rd.,  London,  EC1M  5NN,  UK. 


September  19-20,  1994:  Drug  Discovery  & Commercial 
Opportunities  in  Medicinal  Plants,  Arlington,  VA. 
Contact:  IBC  USA  Conferences  Inc.,  225  Turnpike  Rd., 
Southborough,  MA  01772-1749.  PHN:  (508)  481-6400, 
FAX:  (508)  481-7911. 


September  19-22,  1994:  John  Innes  Symposium:  Biochem- 
istry of  Development,  Norwich,  England.  Contact:  John 
Innes  Centre,  Norwich  Research  Park,  Colney,  Norwich, 
Norfolk,  England  NR4  7UH.  PHN:  44  603  52571,  FAX: 

44  603  56844. 


September  25-27,  1994:  Harnessing  Apomixis:  A New 
Frontier  in  Plant  Science,  College  Station,  TX.  Contact: 
Dr.  David  M.  Stelly,  Dept,  of  Soil  and  Crop  Sciences, 
Texas  A&M  University,  College  Station,  TX  77843-2474. 
PHN:  (409)  845-2745,  FAX:  (409)  862-4733,  EMAIL: 
monosom@rigel.tamu.edu. 

September  30-October  5,  1994:  Structural  Molecular 
Biology  Conference,  Mont  St.  Odile,  France.  Contact: 
Dr.  Josip  Hendekovic,  European  Science  Foundation,  1 
Quai  Lezay-Marnesia,  F-67080  Strasbourge,  Cedex, 
France.  PHN:  (88)  76  71  35,  FAX:  (88)  36  69  87,  TELEX: 
890440. 


October  2-6,  1994:  22nd  Aharon  Katzir-Katchalsky 
Conference:  Plant  Molecular  Biology— Potential 
Impact  on  Agriculture  and  the  Environment,  Koln, 
Germany.  Contact:  Secretariat  22nd  AKK  Conference, 
Aharon  Katzir-Katchalsky  Center,  Weizmann  Institute 
of  Science,  Rehovot  76100,  Israel.  PHN:  972-8-342148, 
FAX:  972-8-474425. 


October  2-6,  1994:  1994  Second  International  Symposium 
on  the  Applications  of  Biotechnology  to  Tree  Culture, 
Protection,  and  Utilization,  Minneapolis,  MN.  Contact: 
Edith  Franson,  Executive  Secretary,  Tree  Biotechnology 
Symposium,  Forestry  Sciences  Laboratory,  P.O.  Box  898, 
Rhinelander,  WI  54501.  PHN:  (715)  362-7474,  FAX:  (715) 
362-7816. 


October  7-10,  1994:  Genetic  & Biochemical  Approaches  for 
Studying  Cell  Death,  American  Society  for  Biochemis- 
try and  Molecular  Biology  Fall  Symposia  1, 
Granlibakken,  Lake  Tahoe,  CA.  Contact:  ASBMB  Fall 
Symposia  Office,  Room  3206,  9650  Rockville  Pike, 
Bethesda,  MD  20814-3998.  PHN:  (301)  530-7010,  FAX: 
(301)  530-7014. 


October  14-17,  1994:  Mechanisms  of  Regulated  Intracellu- 
lar Protein  Degradation:  American  Society  for  Bio- 
chemistry and  Molecular  Biology  Fall  Symposia  2, 
Whistler,  British  Columbia,  Canada.  Contact:  ASBMB 
Fall  Symposia  Office,  Room  3206,  9650  Rockville  Pike, 
Bethesda,  MD  20814-3998.  PHN:  (301)  530-7010,  FAX: 
(301)  530-7014. 


October  16-21,  1994:  Recombinant  DNA  Biotechnology  III 
Conference,  Deauville,  France.  Contact:  Engineering 
Foundation,  Room  303,  245  East  47th  St.,  New  York,  NY 
10017.  PHN:  (212)  705-7837,  FAX:  (212)  705-7441. 

October  28-31,  1994:  Oligonucleotide  Selection  and 

Molecular  Diversity:  American  Society  for  Biochemis- 
try and  Molecular  Biology  Fall  Symposia  3, 
Granlibakken,  Lake  Tahoe,  CA.  Contact:  ASBMB  Fall 
Symposia  Office,  Room  3206,  9650  Rockville  Pike, 
Bethesda,  MD  20814-3998.  PHN:  (301)  530-7010,  FAX: 
(301)  530-7014. 

November  14, 1994:  Cucurbitaceae  94:  Evaluation  and  Enhance- 
ment of  Cucurbit  Germplasm,  South  Padre  Island,  TX.  Contact: 
Dr.  James  R.  Dunlap,  Texas  Agricultural  Experiment  Station,  2415 
East  Highway  83,  Weslaco  TX  78596.  PHN:  (210)  968-5585,  FAX: 
(210)  968-0641,  EMAIL:  j-dunlap@tamu.edu 


28 


Probe 


Volume  4,  No.1/2 


November  13-16,  1994:  Third  International  Symposium  on 
the  Biosafety  Results  of  Field  Tests  of  Genetically 
Modified  Plants  and  Organisms,  Monterey,  CA.  Con- 
tact: Ms.  Pat  Day,  University  of  California,  DANR,  300 
Lakeside  Dr.,  6th  Fir.,  Oakland,  CA  94612-3560  OR 
USDA,  Office  of  Agricultural  Biotechnology.  PHN:  (703) 
235-4419,  FAX:  (703)  235-4429. 

November  17-19,  1994:  1994  San  Diego  Conference:  The 
Genetic  Revolution,  San  Diego,  CA.  Contact:  Scherago 
International,  11  Penn  Plaza,  Suite  1003,  New  York,  NY 
10001.  PHN:  (212)  643-1750,  FAX:  (212)  643-1758, 

EMAIL:  Scherago@Biotech.Net.Com. 

November  21-24,  1994:  Brighton  Crop  Protection  Confer- 
ence: Pest  and  Diseases,  Brighton,  UK.  Contact:  Confer- 
ence Associates  and  Services  Ltd.,  55  New  Cavendish  St., 
London  W1M  7RE,  UK. 


Workshops  and  Courses 

September  26-30  or  October  31-November  4,  1994:  Poly- 
merase Chain  Reaction  Methodology  Workshop, 
Columbia,  MD.  Contact:  Exon-Intron,  Suite  130,  9151 
Rumsey  Rd„  Columbia,  MD  21045-1929.  PHN:  (301)  730- 
3984,  FAX:  (301)  730-3983. 

October  3-7,  1994:  RNA  Isolation  & Characterization 
Workshop,  Columbia,  MD.  Contact:  Exon-Intron,  Suite 
130,  9151  Rumsey  Rd.,  Columbia,  MD  21045-1929.  PHN: 
(301)  730-3984,  FAX:  (301)  730-3983. 

October  10-14,  1994:  Advanced  Course  on  Molecular 
Biology  Workshop,  Leiden,  Netherlands.  Contact:  Dr. 
L.A.  van  der  Meer-Lerk,  Institute  of  Biotechnology 
Studies  Delft,  Kluyver  Laboratory,  Julianalaan  67,  2628 
BC,  Delft,  Netherlands.  PHN:  015-78  51  40,  FAX:  015-78 
23  55. 

October  17-20,  1994:  Polymerase  Chain  Reaction  Tech- 
niques and  DNA  Sequencing  Lecture  Course,  Lake 
Tahoe,  NV.  Contact:  Director,  Center  for  Advanced 
Training  in  Cell  and  Molecular  Biology,  Catholic  Univer- 
sity of  America,  620  Michigan  Ave.,  NE,  Washington,  DC 
20064.  PHN:  (202)  319-6161,  FAX:  (202)  319-4467, 

EMAIL:  millerm@cua.edu. 

October  17-20,  1994:  Recombinant  DNA  Methodology  and 
DNA  Sequencing  Lecture  Course,  Lake  Tahoe,  NV. 
Contact:  Director,  Center  for  Advanced  Training  in  Cell 


and  Molecular  Biology,  Catholic  University  of  America, 
620  Michigan  Ave.,  NE,  Washington,  DC  20064.  PHN: 
(202)  319-6161,  FAX:  (202)  319-4467,  EMAIL: 
millerm@cua.edu. 

December  14-17,  1994:  International  Symposium  on  Plant 
Molecular  Biology  and  Biotechnology  Workshop,  New 
Delhi,  India.  Contact:  G.  Chatterjee,  International  Centre 
for  Genetic  Engineering  and  Biotechnology,  Aruna  Asaf 
Ali  Marg,  New  Delhi  110067,  India.  PHN:  (Oil)  6867356, 
FAX:  (011)  6862316. 

January  7-13,  1995:  Plant  Cell  Biology:  Mechanisms, 

Molecular  Machinery,  Signals,  and  Pathways:  Keystone 
Symposium,  Taos,  NM.  Contact:  Keystone  Symposia, 
Drawer  1630,  Silverthorne,  CO  80498.  PHN:  (303)  262- 
1230,  FAX:  (303)  262-1525. 


Future  Events 


January  15-19,  1995:  Plant  Genome  III,  San  Diego,  CA. 
Contact:  Plant  Genome  III,  c/o  Scherago  International 
Inc.,  11  Penn  Plaza,  New  York,  NY  10001.  PHN:  (212) 
643-1750,  FAX:  (212)  643-1758,  EMAIL: 
scherago@biotechnet.com 

February  4-9,  1995:  Advances  in  Gene  Technology:  Protein 
Engineering  and  Structural  Biology:  Miami  Bio/ 
Technology  Winter  Symposium,  Ft.  Lauderdale,  FL. 
Contact:  Miami  Bio/Technology  Winter  Symposia,  P.O. 
Box  016129  (M823),  Miami,  FL  33101.  PHN:  (800)  642- 
4363,  FAX:  (305)  324-5665,  EMAIL: 
mbws@mednet.med.miami.edu 

March  5-9,  1995:  XVIII  Eucarpia  Symposium:  Ornamental 
Plant  Improvement,  Classical  and  Molecular  Ap- 
proaches, Tel  Aviv,  Israel.  Contact:  Dan  Knassim  Ltd., 
P.O.  Box  57005,  Tel  Aviv,  61570  Israel.  PF1N:  (972)  3- 
5626470,  FAX:  (972)  3-5612303. 

April  23-27,  1995:  3rd  International  Union  of  Biochemistry 
and  Molecular  Biology  Conference:  Molecular  Recogni- 
tion, Singapore.  Contact:  3rd  IUBMB  Conference  Coordi- 
nator, Ken-Air  Destination  Management  Company,  35 
Selegie  Rd.,  09-19  Parklane  Shopping  Mall,  Singapore 
0718.  PHN:  (65)  336-8857/8,  FAX:  (65)  336-3613. 

May  13-17,  1995:  Ninth  International  Biotechnology 
Meeting  & Exhibition,  San  Francisco,  CA.  Contact: 
Biotechnology  Industry  Organization,  1625  K St.,  NW, 


Probe 


July  1993  -July  1994 


Suite  1100,  Washington,  DC  20006-1604.  PHN:  (202)  857- 
0244,  FAX:  (202)  331-8132  or  (202)  857-0237. 


July  4-7,  1995:  9th  International  Rapeseed  Congress, 

Cambridge,  England.  Contact:  Denis  Kimber,  44  Church 
St.,  Haslingfield,  Cambridge,  CB3  7JE,  England. 

July  14-19,  1995:  15th  International  Conference  on  Plant 
Growth  Substances,  Minneapolis,  MN.  Contact:  Gary 
Gardner,  Dept,  of  Horticultural  Science,  University  of 
Minnesota,  305  Alderman  Hall,  St.  Paul,  MN  55108.  FAX: 
(612)  624-3606,  EMAIL:  ggardner@maroon.tc.umn.edu 


August  6-11,  1995:  10th  International  Workshop  on  Plant 
Membrane  Biology,  Regensburg,  Germany.  Contact: 
Widmar  Tanner,  Lehrstuhl  fur  Zellbiologie  und 
Pflanzenphysiologie,  Universitat  Regensburg, 
Universitatsstrasse  31,  93053  Regensburg,  Germany. 
FAX:  49-943-3352. 


August  6-12,  1995:  20th  World  Congress  of  the  Interna- 
tional Union  of  Forestry  Research  Organisations, 
Tampere,  Finland.  Contact:  Professor  Risto  Seppala, 
Finnish  Forest  Research  Institute,  IUFRO-95,  Secretariat 
Unioninkatu  40A  00170,  Helsinki,  Finland. 


Harnessing 

Apomixis: 

A New 


Speakers  and  posters  will  cover 
various  genetic,  molecular,  physi- 
ological, cytological  and  evolution- 
ary aspects  of  asexual  reproduction 
through  seed  and  its  application  to 
crop  improvement. 


Frontier  in 
Plant  Science 


September  25-27 


Hilton  Hotel  and 
Conference  Center 
College  Station 


Contact: 


Related  topics  in  plant  sexual 
reproduction  also  will  be  pre- 
sented. Some  financial  support 
for  international  attendees  will 
be  available. 


Dr.  David  M.  Stelly 
Department  of  Soil  and  Crop  Sciences 
Texas  A&M  University 
College  Station,  Texas  77843-2474 
E-mail:  monosom@rigel.tamu.edu 
Phone:  (409)  845-2745 
Fax:  (409)  862-4733 


Probe 


Volume  4,  No.1/2 


Survey  of  Synonymous  Codon  Usage 
in  Nuclear  Genes  of  Arabidopsis, 
Soybean  and  Maize 

Julia  Bailey-Serres  and  Sheila  L.  Fennoy 

Department  of  Botany  and  Plant  Sciences , University  of  California 
Riverside , CA  92521-0124 


The  overall  bias  in  synonymous 
codon  usage  of  a genome  is  species- 
specific.  Analysis  of  protein  coding 
regions  of  small  samples  of  plant 
genes  for  a number  of  species  re- 
vealed codon  usage  biases.1  The 
synonymous  codon  usage  of  nuclear 
genes  of  plants  varies  mainly  in  the 
bias  toward  C or  G versus  A or  U in  the  silent  third 
nucleotide  position.  Nuclear  gene  coding  regions  of 
monocots  are  enriched  in  codons  ending  in  C and  G, 
whereas  dicots  have  a higher  frequency  of  codons  ending 
in  A and  U. 

We  used  a multivariate  statistical  analysis  to 
examine  codon  usage  in  maize.  More  biased  codon  usage 
was  recognized  among  more  highly  expressed  genes, 
whereas  more  random  codon  usage  was  observed  among 
more  lowly  expressed  genes.  Our  work  indicates  that  the 
overall  codon  usage  patterns  in  maize  reflect  the  G+C 
content  of  the  genome.  Codon  usage  bias  of  individual 
genes  may  not  solely  reflect  the  nucleotide 
compositional  bias  of  a chromosomal 
region,  but  may  be  affected  by  selection  on 
the  silent  third  nucleotide.2 

The  accumulation  of  DNA  sequence 
data  for  a large  number  of  nuclear  genes  of 
plants  provided  an  opportunity  to  further 
examine  synonymous  codon  usage.  Table  1 shows  a 
summary  of  codon  usage  for  three  plant  species,  maize 
( Zea  mays  L.),  soybean  ( Glycine  max  L.),  and  Arabidopsis 
(. Arabidopsis  thaliana).  Non-duplicate  protein  coding 


sequences  were  obtained  from  the  September  1992 
releases  of  GenBank  and  EMBL  databases  and  the 
literature,  and  the  relative  synonymous  codon  usage  was 
determined.  The  synonymous  codons  used  at  a higher 
frequency  in  these  data  sets  are  indicated  with  an  aster- 
isk. 

Information  on  codon  usage  is  useful  for  the  design 
of  degenerate  oligonucleotide  primers  for  PCR  amplifica- 
tion of  regions  encoding  conserved 
proteins.  In  addition,  consideration  of 
G+C  content  or  codon  usage  appears 
to  be  important  for  high  levels  of 
expression  of  bacterial  genes  in 
plants.34  Further  systematic  analyses 
are  needed  to  determine  the  role  of  the  G+C  content  and 
codon  usage  in  regulating  gene  expression. 

References 

1.  Campbell,  W.H.  and  Gowri,  G.  (1990)  Plant  Physiol,  92, 
1-11. 

2.  Fennoy,  S.L.  and  Bailey-Serres,  J.  (1993)  Nucl.  Acids  Res., 
21,  5294-5300. 

3.  Perlak,  F.J.,  Fuchs,  R.L.,  Dean,  D.A.,  McPherson,  S.L.  and 
Fischhoff,  D.A.  (1991)  Proc.  Natl.  Acad.  Sci.  USA,  88, 
3324-3328. 

4.  Koziel,  M.G.,  Beland,  G.L.,  Bowman,  C.,  Carozzi,  N.B., 
Crenshaw,  R.,  Crossland,  L.,  Dawson,  J.,  Desai,  N.,  Hill, 
M.,  Kadwell,  S.,  Launis,  K.,  Lewis,  K.,  Maddox,  D., 
McPherson,  K.,  Meghji,  M.R.,  Merlin,  E.,  Rhodes, 

R., Warren,  G.W.,  Wright,  M.  and  Evola,  S.V.  (1993)  Bio/ 
Technology, 11,  194-200. 


Probe 


July  1993 -July  1994 


31 


Table  1 


Malza 

Soybean 

Arabidopsis 

AA 

CODON 

N 

RSCU 

N 

RSCU 

N 

RSCU 

Ala 

GCU 

841 

0.92 

606 

1.5* 

1 572 

1 .87* 

GOC 

1319 

1 .45* 

376 

0 93 

653 

0 78 

GCA 

511 

0.56 

492 

1 .22 

728 

0.87 

<333 

955 

1.05 

1 42 

0.35 

403 

0.48 

Leu 

UUA 

102 

0.19 

215 

0.65 

338 

0.62 

UUG 

407 

0.75 

474 

1.44 

860 

1.36 

cuu 

527 

0.97 

531 

1 .62* 

1024 

1 .62* 

cue 

993 

1 .83* 

362 

1.1 

794 

1 .26 

CUA 

220 

0.41 

161 

0.49 

359 

0.57 

CUG 

1000 

1 .85* 

229 

0 7 

359 

0 57 

Gly 

G3U 

640 

0 83 

508 

1 .29* 

1287 

1 5* 

GOC 

1417 

1 .83* 

293 

0.75 

458 

0.53 

GGA 

476 

0.61 

498 

1 .27* 

1287 

1.5* 

(333 

566 

0.73 

274 

0.7 

396 

0 46 

Val 

GUU 

512 

0.77 

634 

1.56* 

1172 

1 58* 

GUC 

883 

1 33 

243 

0.6 

712 

0 96 

GUA 

1 99 

0.30 

1 82 

0.45 

269 

0.36 

GLG 

1062 

1.6* 

568 

1.4 

805 

1.09 

Ser 

AGU 

1 82 

0 43 

420 

1 .38* 

487 

0.93 

AGC 

640 

1 .54* 

254 

0.84 

480 

0.92 

ICU 

353 

0.85 

354 

0.65 

835 

1 .6* 

UOC 

647 

1 .55* 

1 14 

0.38 

4 59 

0.88 

UCA 

300 

0.72 

337 

1.11 

576 

1 .1 

LCG 

376 

0 90 

345 

1.13 

296 

0.57 

Pro 

ecu 

459 

0.84 

586 

1.21 

730 

1 .44* 

OOC 

581 

1 .06 

325 

0.7 

278 

0.55 

CCA 

455 

0.83 

899 

1 .85* 

707 

1 .40* 

COG 

701 

1 28* 

131 

0.27 

311 

0 61 

Glu 

GAA 

555 

0.49 

854 

1 .00* 

1219 

0.91  * 

GAG 

1724 

1.51  * 

875 

1.01* 

1465 

1 .09* 

Arq 

03U 

231 

0.66 

178 

0.98 

433 

1.22 

CGC 

643 

1 .85* 

153 

0.84 

1 52 

0.43 

CGA 

125 

0 36 

108 

0.59 

1 95 

0.55 

OGG 

314 

0.90 

51 

0 28 

1 39 

0.39 

AGA 

207 

0 59 

320 

1 .76* 

663 

1 .87* 

AGG 

569 

1 63* 

284 

1.56* 

544 

1 54* 

Thr 

ACU 

369 

0.76 

421 

1.4* 

827 

1 .39* 

ACC 

777 

1 .61  * 

330 

1.1 

599 

1 01 

ACA 

328 

0.68 

351 

117 

645 

1.08 

AGG 

459 

0.95 

101 

0.34 

310 

0 52 

Lys 

AAA 

370 

0.39 

629 

0 8 

1043 

0.78 

AAG 

1 540 

1 61  * 

952 

1 .20“ 

1635 

1 22* 

Asp 

GAU 

645 

0.68 

716 

1 .22* 

716 

1.22* 

GAG 

1240 

1 .31  * 

453 

0.78 

453 

0.78 

lie 

AUU 

442 

0.81 

571 

1 43* 

998 

1.24* 

AUC 

1013 

1 .86* 

353 

0 88 

1035 

1 .29* 

AUA 

181 

0.33 

278 

0.69 

382 

0.47 

Gin 

CAA 

465 

0.61 

586 

1.16* 

763 

0.98* 

CAG 

1066 

1 .39* 

422 

0.84 

789 

1 .02* 

Phe 

UUU 

332 

0.50 

470 

0.98* 

682 

0.79 

UUC 

1006 

1 .5* 

486 

1 .02* 

1039 

1 .21  * 

Asn 

AAU 

353 

0 55 

497 

0.91 

719 

0 82 

AAC 

923 

1 .44* 

599 

1 .09* 

1043 

1.18* 

Tyr 

UAU 

236 

0.45 

421 

0.94* 

455 

0.76 

UAC 

808 

1 55* 

473 

1 .06* 

745 

1 .24* 

His 

CAU 

265 

0.63 

301 

1.11* 

413 

1.00* 

CAC 

572 

1 .37* 

243 

0 89 

411 

1 .00* 

Met 

AUG 

891 

1.00 

505 

1 .00 

1 1 06 

1 .00 

Cys 

UGU 

151 

0 49 

1 33 

0 81 

332 

1 .04* 

UGC 

470 

1 .51* 

1 97 

1.19* 

307 

0.96 

Trp 

LDG 

420 

1 .00 

262 

1 .00 

471 

1 .00 

TER 

UGA 

45 

1 .35* 

23 

0 99 

43 

1 .15* 

UAA 

23 

0.69 

31 

1.29* 

43 

1.15* 

LAG 

32 

0.96 

1 7 

0.72 

26 

0.7 

Summary  of  relative  synonymous  codon  usage  in  three  plant  species. 
Codon  usage  was  tabulated  (N)  for  maize  (100  genes),  soybean  (71 
genes),  and  Arabidopsis  (1 12  genes).  The  relative  synonymous 
codon  usage  (RSCU)  is  the  observed  frequency  divided  by  the 
expected  frequency  assuming  random  codon  usage.  The  most 
frequently  used  synonym  for  each  amino  acid  of  each  plant  is  marked 
by  an  asterisk. 


Register  Today ... 

Early  registration  is  encouraged 
to  guarantee  a place  at  the 
Third  International  Symposium 
on  the  Biosafety  Results  of  Field 
Tests  of  Genetically  Modified 
Plants  and  Microorganisms. 

The  symposium  will  be  held 
November  13-16,  1994,  in 
Monterey,  CA 

Seven  panels  will  convene  to 
explore  such  questions  as: 

• Can  small-scale  results  be 
extrapolated  in  assessing  risk? 


• Are  there  unique  risks  for 
Centers  of  Diversity? 

• Can  new  viral  pathogens  be 
generated  from  transgenic 
plants? 


Experiences  in  the  commercial- 
ization of  transgenic  crop  plants 
also  will  be  discussed. 

To  receive  a program  announce- 
ment and  a complete  agenda, 
please  fax  your  request  to  Office 
of  Agricultural  Biotechnology, 


(703)  235-4429. 


[•] 


Probe 


Volume  4,  No.1/2 


Plant  Genome  Analysis  by  Single 
Arbitrary  Primer  Amplification 


Peter  M.  Gresshoff 
Plant  Molecular  Genetics 

Center  for  Legume  Research  and  Institute  of  Agriculture 
The  University  of  Tennessee 
Knoxville , TN  37901-1071 


Molecular  genetics 
approaches  have 
enriched  the  resolu- 
tion of  plant  genome 
analysis.  The  abihty  to  clone  and 
sequence  specific  genome  regions  has 
added  sequence-based  information 
to  our  understanding  of  plant 
genomes  derived  from  cytogenetics 
and  large-scale  DNA  analyses  (such 
as  reassociation  analysis). 

While  the  database  of  DNA 
sequences  is  exponentially  growing, 
methods  are  needed  to  investigate 
plant  genomes  at  a level  of  complex- 
ity above  the  primary  sequence,  but 
below  the  cytogenetic,  karyotypic 
arrangement. 

Single,  arbitrary  primer-based 
DNA  amplification  techniques  (DAF, 
RAPD  and  AP-PCR)  were  developed 
(Caetano-Anolles  et  al.,  1991a; 
Williams  et  al.,  1990;  Welsh  and 
McClelland,  1990),  extending  the 
utility  of  PCR  to  general  genome 
analysis  (fig.  1).  Because  of  a plethora 
of  terms,  we  proposed  the  general 
acronym  MAAP  (Multiple  Arbitrary 


Amplicon  Profiling;  Caetano-Anolles 
et  al.,  1992b,  1993,  1994). 

In  essence,  MAAP  involves  the 
use  of  a short,  arbitrarily  chosen 
ohgonucleotide  primer  which, 
annealed  to  DNA,  will  direct  DNA 
amplification  of  multiple  genome 
regions  (amplicons;  Mullis,  1991). 
Temperature  cycling  and  the  use  of  a 
thermostable  DNA  polymerase  are 
common  components  with  the  more 
specific  and  targeted  PCR.  In  con- 
trast to  PCR,  MAAP  procedures  use 


Figure  1:  Uses  of  single  primer 
amplification  methods. 

a single  primer  which  is  of  arbitrary 
sequence.  MAAP  intentionally 
generates  multiple  products,  which 


itself  would  be  a rather  undesirable 
result  in  a PCR  reaction.  MAAP  is 
general,  so  that  a primer  used  for  one 
species  can  be  used  repeatedly  for 
others,  even  if  evolutionary  distances 
between  the  template  DNAs  are 
large. 

Amplification  products  are 
separated  and  recorded  by  a variety 
of  detection  methods;  in  all  cases,  a 
linear  array  of  signals  generates  a 
profile,  which  is  representative  for 
the  target  DNA  and  specified  by  the 
DNA  sequence  of  the  primer.  Varia- 
tions in  primer  sites  on  the  target 
DNA,  length  variations  between 
primer  sites,  and  possibly  changes  in 
the  secondary  structure  of  target 
DNA  between  or  flanking  the  primer 
recognition  sites,  generate  molecular 
polymorphisms.  These  amplification 
polymorphisms  define  molecular 
regions  of  the  plant  genome  and  thus 
can  be  used  as  (1)  potential  sequence 
tagged  sites  for  positional  cloning 
approaches,  or  (2)  components  of 
profile  used  in  DNA  profiling  and 
diagnostics. 


Abbreviations:  PCR=polymerase  chain  reaction;  DAF=DNA  amplification  fingerprinting;  AP.-PCR=arbitrary  primer-PCR;  MAAP=multiple  arbitrary 
amplicon  profiling;  nf=nucleotide;  bp= base  pair;  PAG£=polyacrylamide  gel  electrophoresis;  RAPD=random  amplified  polymorphic  DNA; 
RFLP=restriction  fragment  length  polymorphism 


July  1993  -July  1994 


Probe 


33 


Three  Techniques 

MAAP  procedures  were  developed 
independently,  and  apparently 
concurrently,  in  three  laboratories. 
Welsh  and  McClelland  (1990)  devel- 
oped AP-PCR,  which  uses  PCR- 
length  primers  [18  to  32  nt]  of 
arbitrary  sequence  to  amplify  target 
DNA  under  low  stringency  anneal- 
ing conditions  for  two  amplification 
cycles.  This  allows  abundant  mis- 
matching and  the  generation  of 
multiple  amplification  products 
(equivalent  to  a PCR  reaction  having 
gone  wrong).  Increased  stringency  of 
annealing  at  later  amplification 
cycles  generated  reproducible 
products  which  were  resolved  on 
polyacrylamide  gels  and  detected 
by  autoradiogra- 
phy. 

Williams  et 
al  (1990)  invented 
the  RAPD  proce- 
dure, in  which  an  arbitrary  primer 
of  either  9 or  10  nt  produced  amplifi- 
cation products  after  temperature 
cycling.  RAPD  products  are  rou- 
tinely resolved  on  agarose  gels  and 
visualized  by  ethidium  bromide. 

This  provides  a rapid  method  of 
scanning  a genome.  Alternative 
methods  of  detection,  such  as  PAGE 
and  silver-staining,  coupled  with 
careful  optimization  of  amplification 
parameters  (Collins  and  Symons, 
1993)  improved  the  utility  of  the 
approach.  RAPD  is  widely  used 
because  of  its  simplicity  and  low-cost 
instrumentation. 

Caetano-Anolles  et  al.  (1991a,b) 
developed  DNA  Amplification 
Fingerprinting  (DAF).  Of  all  MAAP 
procedures,  DAF  utilizes  the  shortest 
primers,  down  to  5 nt  in  length.  The 


optimal  length  was  found  to  be  8 nt, 
a length  which  does  not  produce 
efficient  amplification  with  RAPD. 
Informative  amplification  profiles 
were  generated  with  5 nt  primers  (5- 
mers),  using  soybean  DNA  as  a 
template  (Caetano-Anolles  et  al., 
1993). 

DAF  products  are  routinely 
separated  by  thin  polyacrylamide 
gels,  backed  onto  plastic  Gel-Bond 
film.  This  gel-plastic  support,  which 
provides  support  during  the  washing 
steps  and  helps  preserve  the  original 
gel,  is  stained  by  an  improved  silver- 
staining  method  (Bassam  et  al.,  1991; 
Caetano-Anolles  and  Gresshoff, 
1994a),  which  detects  DNA  at  about 


Developments 


1 pg  mirr2.  Resultant  gels  are  air- 
dried  and  kept  for  permanent  record 
and  evaluation. 

Pattern  Detection 

The  PAGE/ silver-staining  technique 
provides  a low-cost,  high-throughput 
analytical  method  of  DAF  products. 
DAF  products  were  also  resolved  by 
alternative  methods.  Agarose  gels 
give  clear  resolution,  but  fewer 
products  (Prabhu  and  Gresshoff, 
1994).  Fluorochrome  labeled  octamer 
primers  were  generated  which  then 
directed  amplification  of  plant  DNA 
(Caetano-Anolles  et  al.,  1992a).  The 
resultant  amplification  products 
were  separated  on  an  ABI  Sequencer 
using  Gene  Scanner  software.  Single 
nucleotide  resolution  was  obtained 


for  lower  sized  amplification  prod- 
ucts. Tests  using  capillary  electro- 
phoresis have  been  promising  (Dr. 
Patrick  Williams,  DNA  Testing 
Faboratory,  AFIP,  Gaithersburg,  MD; 
personal  communication),  providing 
separation  of  single  samples  in  30 
minutes.  In  general,  DAF  generates 
scoreable  polymorphisms  in  the 
molecular  size  range  from  100  to  800 
bp.  Recently,  we  have  used  the  pre- 
cast and  automated  PhastGel  system 
(Pharmacia  Inc.)  to  obtain  profiles  for 
pathogenic  nematodes  on  soybean 
(Baum  et  al.,  1994).  Bands  at  higher 
molecular  weight  (up  to  1500  bp) 
were  scoreable;  species  and  race- 
specific  polymorphisms  were  de- 
tected. Denaturing  gradient  gel 
electrophoresis 
(DGGE)  is  another 
method  which 
would  help  to 
distinguish 

polymorphic  products  of  wheat 
(He  et  al.,  1992). 

Genetic  Uses  of  DAF 

The  ability  to  detect  molecular 
markers  closely  associated  with 
genes  of  agricultural  importance 
makes  marker-based  breeding  an 
attractive  proposition.  The  need  for 
maintaining  large  plant  populations 
through  advanced  breeding  cycles 
can  be  reduced  by  detecting  het- 
erozygotes. MAAP  markers  con- 
verted through  cloning,  partial 
sequence  analysis  and  specific  PCR 
primer  synthesis  may  provide 
SCARs  (sequence  characterized 
amplified  regions),  which  are  diag- 
nostic for  either  a gene  region  in  a 
plant  or  a pathogen.  Figure  2 car- 
toons the  utility  of  RFFPs  and  MAAP 


34 


Probe 


Volume  4,  No.1/2 


DIAGNOSTIC  PCR  PRODUCT 


Figure  2:  RFLP  and  MAAP  markers  used 
as  diagnostic  tools  for  genome  analysis. 
Partial  sequencing  of  clones,  which  are 
linked  to  your  favorite  gene  (yfg!) 
provides  information  for  specific  PCR 
primers,  ivhich  in  turn  generate  a 
diagnostic  product.  A sequence  character- 
ized amplified  region  (SCAR)  was 
demonstrated  for  the  supernodulation 
gene  of  soybean  by  Kolchinsky  et.  al. 

(1994) 

markers  in  generating  diagnostic 
tools.  For  example,  it  may  be  possible 
to  find  markers  specific  for  a soybean 
nematode  race  (see  Baum  et  al., 

1994),  to  convert  it  to  a SCAR,  then 
use  a diagnostic,  proactive  test  on 
agricultural  soil  to  predict  which 
nematode  race  is  predominant  in  the 
field  prior  to  planting. 

The  ability  to  generate  many 
amplification  products  means  that 
DAF  is  very  efficient  in  scanning  the 
genome  of  an  organism  for  variable 
sites.  In  a survey  of  25  primers  (all 
octamers),  Prabhu  and  Gresshoff 
(1994),  working  with  G.  max  and  G. 
soja,  detected  an  average  of  1.5 
AFLPs  per  primer.  Interestingly, 
RAPD  gels  of  soybean  produce  an 
average  of  5 to  7 scoreable  bands, 
while  DAF  in  soybean  produced  an 
average  of  20  to  25  bands.  Accord- 


ingly, the  ratio  of  scored  polymor- 
phism to  scoreable  band  is  nearly  the 
same,  that  DAF  is  not  picking  up 
more  AFLPs  because  of  the  shorter 
primer  length,  but  because  of  the 
detection  method. 

DAF  markers  were  shown  to  be 
repeatable  polymorphisms  in  differ- 
ent DNA  isolations,  operators,  time 
periods,  and  amplifications.  They  are 
heritable,  as  are  about  75%  of  AFLPs 
between  G.  max  and  G.  soja  segre- 
gated as  dominant  Mendelian 
markers  in  F2  populations  (Prabhu 
and  Gresshoff,  1994;  Caetano-Anolles 
et  al.,  1993).  Interestingly,  the  other 
25%  segregated  in  a uniparental  way, 
being  either  maternal  or  paternal. 
Maternal  inheritance  presumably 
stems  from  amplification  of  cytoplas- 
mic replicons.  As  yet,  paternal 
replication  is  unexplained,  and  may 
represent  either  highly  repeated 
chromosomal  replicons  or  possibly 
alterations  from  normal  cytoplasmic 
inheritance  patterns  in  soybean. 

Recombinant  Inbred  Lines 

Several  DAF  polymorphisms  were 
mapped  in  recombinant  inbred  lines 
of  soybean  (Prabhu  and  Gresshoff, 
1994).  The  use  of  inbred  lines  is  very 


convenient  for  DAF,  as  the  hnes  are 
predominantly  homozygous.  Since 
DAF  markers  are  dominant,  it  is 
impossible  to  distinguish  the  domi- 
nant homozygote  from  the  heterozy- 
gote. Accordingly,  in  normal  F2 
populations,  larger  sample  numbers 
are  required  to  obtain  data  equiva- 
lent to  data  obtained  from  the 
analysis  of  a codominant  (e.g.,  RFLP) 
marker.  In  recombinant  inbreds, 
however,  DAF  and  RFLP  markers 
share  the  same  statistical  advantages. 
Figure  3 provides  a summary  of 
some  RIL  mapping  data  (conducted 
in  collaboration  with  Dr.  Gordon 
Lark,  Utah). 

The  large  number  of  products 
allows  a high-density  genotyping 
and  genotype  differentiation 
(Gresshoff,  1992).  This  form  of 
fingerprinting  is  similar  to  the 
Universal  Product  Code,  in  which 
bars  and  spaces  define  a product. 
Reliable  exclusion  is  obtained  when 
one  or  more  bands  differ  between 
samples.  Inclusion  is  more  difficult, 
as  many  primers  need  to  be  tested, 
frequency  of  variation  within  the 
sampled  species  needs  to  be  known, 
and  careful  statistical  statements 
need  to  be  generated.  One  cannot 


use  recombinant  inbred  lines  from  Gordon  lark  (Utah) 

^ parents  eulflvass  Mlnsoy  and  Ndr;  advanced  to  FI  1 
) DAF  markers  seyegate  1:1;  absence  of  heterozygotes 
CH  DAF  maikes  are  dominant,  and  may  detect  chloroplast  DNA 

Rllmap:  :.icu  ucu 

K390  K314  K 265  K544a  IA186  A 245a  SCD 

DAF220 

Figure  3:  Summary  of  mapping  of  one  DAF  marker  on  recombinant  inbred 
lines  (Fll)  derived  from  a cross  of  soybean  cultivars  Minsoy  and  Noir. 


July  1993  -July  1994 


Probe 


35 


declare  with  100%  certainty  that  two 
things  are  the  same;  the  statement 
must  always  be  probabilistic.  It  is  up 
to  the  user  (society,  courts,  scien- 
tists) to  concur  on  acceptable  levels 
of  confidence  for  such  probabilities. 

Fingerprint  Applications 

DAF  allowed  the  easy  distinction  of 
variant  turfgrass  material  in  com- 
mercial plots  (Callahan  et  al.,  1993). 
For  example,  foundation  stock  from 
several  geographic  locations  gave 
identical  profiles  for  Bermudagrass 
Tifway  419,  while  samples  analyzed 
from  golf  course  owners  repeatedly 
showed  major  variation.  The  appli- 
cation of  DNA  tests  to  the  turfgrass 
industry  is  a major  challenge  in  an 
area  of  repeated  vegetative  propaga- 
tion, triploidy  and  genetic  instability. 
Using  DAF  markers.  Weaver  (1994) 
developed  a phylogenetic  tree  of 
centipedegrasses. 

Sunflower  material  provided 
by  a seed  company  was  categorized 
into  several  groups.  Some  common 
bands  permitted  the  suggestion  of  a 
possible  pedigree.  This  type  of 
analysis  has  utility  for  product 
verification  and  plant  variety  rights. 

The  determination  of  genetic 
identity  is  also  essential  for  the 
determination  of  plant  product 
quality,  as  many  food  manufacturers 
use  processes  directly  optimized  for 
a specific  biological  feedstock.  This 
industry  relies  on  biological  mate- 
rial; it  is  essential  that  quality  biologi- 
cal feedstock  enters  the  manufactur- 
ing process.  Often  it  is  impossible  to 
inspect  the  source  plant  as  one  looks 
at  a harvested  product.  It  is  for  these 
industrial  and  related  horticultural 
applications  that  a new  technology 


was  needed.  DNA  analysis  has 
provided  an  additional  way  by  which 
closely  related  organisms  are  distin- 
guished for  industrial.,  manufactur- 
ing, and  retailing  purposes. 

DAF  markers  are  useful  in 
defining  closely  linked  regions  in 
bulked  segregant  analysis 
(Michelmore  et  al.,  1991).  The  avail- 
ability of  large  primer  sets  and  the 
generation  of  multiple  amplification 
products  result  in  the  efficient 
screening  of  the  genome. 

Induced  plant  mutations  have 
the  advantage  of  being  in  near- 
isogenic  background  as  the  genetic 
difference  between  parent  and 
mutant  is  minimal.  Using  25  DAF 
primers,  Caetano-Anolles  et  al.  (1993) 
showed  that  the  induced 
supernodulation  mutant  nts382  and 
its  wild-type  parent  cv.  "Bragg"  did 
not  show  polymorphisms  despite  the 
pairwise  comparison  of  nearly  500 
amplification  products.  Only  in  the 
use  of  MAAP,  in  which  the  target 
DNA  was  predigested  with  two 
restriction  nucleases  (four  base 
cutters)  and  then  amplified  with  a 
single  octamer,  could  polymorphisms 
be  detected  between  mutant  and 
wild-type  parent.  Only  19  primers 
were  needed  to  reveal  42  AFLPs. 
Fourteen  of  these  segregated  at  100% 
with  the  supernodulation  phenotype 
in  G.  soja  (wild-type)  and  G.  max 
(mutant  ) derived  F2  populations. 
Some  AFLPs  distinguished  between 
the  nts382  and  ntsl007  alleles.  It  is 
likely  that  these  are  valuable  markers 
close  to  the  tits  locus  and  their  cloning 
and  further  characterization  will 
facilitate  the  isolation  and  ordering  of 
yeast  artificial  chromosomes  (YACs)  in 
that  region. 


Mini-hairpin  Primers 

Funke  and  Kolchinsky  (1994)  demon- 
strated that  stable  YACs  carrying 
soybean  genomic  DNA  can  be 
constructed,  with  an  average  size  of 
about  200  kb  (maximum  900  kb). 
About  7%  represented  chloroplastic 
DNA.  The  combination  of  clustered 
molecular  markers,  the  ability  to 
generate  medium-sized  YAC  clones, 
end-clones  and  possible  contigs, 
increase  the  chances  of  isolating 
soybean  regions  carrying  develop- 
mentally  significant  genes.  Caetano- 
Anolles  and  Gresshoff  (1994b)  used 
mini-hairpin  primers  in  a DAF 
reaction  to  profile  such  soybean 
YACs.  The  mini-hairpin  primers  are 
interesting,  because  they  contain  on 
their  5'  end  a 7 nucleotide  fold-back 
loop  (4  nt  in  stem,  3 nt  in  the  loop). 
The  3'  end  can  be  as  short  as  3 nt, 
allowing  the  generation  of  a small  set 
of  64  primers,  which  are  useful  for 
the  characterization  especially  of 
small  genomes  or  genome  compo- 
nents such  as  plasmids  or  YACs. 

These  findings  show  that  single 
primer  DNA  amplification  analysis 
of  plant  genomes  adds  a further 
genetic  tool  to  construct  high-density 
maps  needed  for  positional  cloning 
and  marker-based  breeding  ap- 
proaches. 

References: 

Baum,  T.  J.,  Gresshoff,  P.M.,  Lewis, 
S.A.  and  Dean,  R.A.  (1994)  Charac- 
terization and  phylogenetic  analysis 
of  four  root-knot  nematode  species 
using  DNA  amplification  finger- 
printing and  automated  polyacryla- 
mide gel  electrophoresis.  MPM1  7: 
39-47. 


36 


Probe 


Volume  4,  No.1/2 


Primer  length:  5 nt  mini- 
mum; 8 nt  optimum 

Primer  3'  end  most  important 
for  specificity  of  reaction 

Primer  concentration:  3pM  for 
8-mer  primer  and  up  to 
30pM  for  5-mer  primer 

2mM  MgCl,  optimum  for 
soybean  genome  and  6mM 
optimum  for  bacterial 
genome 

Taq  polymerase  produces 
good  amplification  results 
for  large  fragments 

Truncated  Stoffel  fragment 
should  be  used  to  amplify 
fragments  in  the  50-200  bp 
range 

Excess  template  DNA  (>25 
ng/25  pi  reaction)  reduces 
intrinsic  amplification 
products 

For  a more  complete  discussion  of 
these  parameters,  please  gopher  to: 
gopher.nalusda.gov.  Select 
Information  Centers  from  the 
menu.  Next  select  Plant  Genome 
Data  and  Information  Center.  If 
you  would  like  a hard  copy  of  the 
paper,  please  contact  the  Plant 
Genome  Data  and  Information 
Center  at  the  address  on  page  3. 


Bassam,  B.J.,  Caetano-Anolles,  G.  and 
Gresshoff,  P.M.  (1991)  A fast  and 
sensitive  silver-staining  for  DNA  in 
polyacrylamide  gels.  Analytical 
Biochemistry  196:  80-83. 

Caetano-Anolles,  G.  and  Gresshoff, 
P.M.  (1994a).  Staining  nucleic  acids 
with  silver:  an  alternative  to 
radioisotopic  and  fluorescent 
labeling.  Promega  Notes  45:  13-18. 

Caetano-Anolles,  G.  and  Gresshoff, 
P.M.  (1994b).  DNA  amplification 
fingerprinting  using  arbitrary  mini- 


hairpin oligonucleotide  primers. 
Bio/Technology  12:  619-623. 

Caetano-Anolles,  G.,  Bassam,  B.J.  and 
Gresshoff,  P.M.  (1991a)  DNA 
amplification  fingerprinting  using 
very  short  arbitrary  oligonucleotide 
primers.  Bio/Technology  9:  553-557. 

Caetano-Anolles,  G.,  Bassam,  B.J.  and 
Gresshoff,  P.M.  (1991b)  DNA 
amplificationfingerprinting:  a 
strategy  for  genome  analysis.  Plant 
Molecular  Biology  Reporter  9:  292- 
305. 

Caetano-Anolles,  G.,  Bassam,  B.J.  and 
Gresshoff,  P.M.  (1992a)  DNA 
amplification  fingerprinting  with 
very  short  primers.  In:  Application  of 
RAPD  Technology  to  Plant  Breeding. 
ed.  M.  Neff.  ASHS,  publ.  (St.  Paul, 
MN).  pp  18-25. 

Caetano-Anolles,  G.,  Bassam,  B.J.  and 
Gresshoff,  P.M.  (1992b)  DNA 
fingerprinting:  MAAPing  out  a 
RAPD  redefinition.  Bio/Technologxi 
10:  937. 

Caetano-Anolles,  G.,  Bassam,  B.J.  and 
Gresshoff,  P.M.  (1993)  Enhanced 
detection  of  polymorphic  DNA  by 
multiple  arbitrary  amplicon  profil- 
ing of  endonuclease  digested  DNA: 
identification  of  markers  linked  to 
the  supernodulation  locus  of 
soybean.  Mol.  Gen.  Genetics  241:  57- 
64. 

Caetano-Anolles,  G.,  Bassam,  B.J.  and 
Gresshoff,  P.M.  (1994)  Multiple 
arbitrary  amplicon  profiling  using 
short  oligonucleotide  primers.  Plant 
Genome  Analysis,  ed.  P.M. 

Gresshoff,  CRC  Press,  Boca  Raton, 
FL,  pp  29-46. 

Callahan,  L.  M.,  Caetano-Anolles,  G., 
Bassam,  B.J.,  Weaver,  K. 

MacKenzie,  A.  and  Gresshoff,  P.M. 
(1993)  DNA  fingerprinting  of 
turfgrass.  Golf  Course  Management. 
June  issue,  pp  80-86. 

Collins,  G.G.  and  Symons,  R.H.  (1993) 
Polymorphisms  in  grapevine  DNA 
detected  by  the  RAPD  PCR  tech- 
nique. Plant  Molecular  Biology 
Reporter  11:  105-112. 

Funke,  R.  P.  and  Kolchinsky,  A.M.  (1994) 
Plant  yeast  artificial  chromosome  libraries 


and  their  use:  status  and  some  strategic 
considerations.  Plant  Genome  Analysis,  ed. 
P.M.  Gresshoff,  CRC  Press,  Boca  Raton, 
FL  pp.  125-134. 

Gresshoff,  P.M.  (1992)  DNA  finger- 
printing brings  high-tech  genetics 
into  commercial  greenhouses. 

Grower  Talks  (July  92),  pp  119-127. 

He,  S.,  Ohm,  H.,  and  McKenzie,  S. 
(1992)  Detection  of  DNA  sequence 
polymorphisms  among  wheat 
varieties.  Theor.  Appl.  Genet.  84:  573- 
578. 

Kolchinsky,  A.,  Landau-Ellis,  D., 
Angermuller,  S.,  Deckert,  J.,  and 
Gresshoff,  P.M.  (1994)  Molecular 
analysis  of  a polymorphic  DNA 
region  tightly  linked  to  the 
supernodulation  (nts)  locus  of 
soybean  (in  review). 

Michelmore,  R.W.,  Paran,  I.  and 

Kesseli,  R.V.  (1991)  Identification  of 
markers  linked  to  disease  resistance 
genes  by  bulked  segregant  analysis: 
a rapid  method  to  detect  markers  in 
specific  genomic  regions  using 
segregating  populations.  Proc.  Natl. 
Acad.  Sci.  (USA)  88:  9828-9832. 

Mullis,  K.B.  (1991)  The  polymerase 
chain  reaction  in  an  anemic  mode: 
how  to  avoid  cold 
oligodioxyribonuclear  fusions.  PCR 
Meth.  Applic.  1:  1-4. 

Prabhu,  R.R.  and  Gresshoff,  P.M. 

(1994)  Inheritance  of  polymorphic 
markers  generated  by  DNA  amplifi- 
cation fingerprinting  and  their  use 
as  genetic  markers  in  soybean.  Plant 
Molecular  Biology  (in  press). 

Weaver,  K.  (1994)  MS  dissertation. 

The  University  of  Tennessee, 
Knoxville. 

Welsh,  J.  and  McClelland,  M.  (1991) 
Fingerprinting  genomes  using  PCR 
with  arbitrary  primers.  Nucleic 
Acids  Res.  18:  7213-7218. 

Williams,  J.G.K,  Kubelik,  A.R,  Livak,  K.J, 
Rafalski,  J.A.  and  Tingey,  S.V.  (1990) 
DNA  polymorphisms  amplified  by 
arbitrary  primers  are  useful  genetic 
markers.  Nucleic  Acids  Res.  18  6531-6535. 

♦ 


Probe 


July  1993 -July  1994  37 


Other  Pursuits 

Distinctive  Biology  of  Forest  Trees 
Highlighted  at  Sixth  International 

Meeting 

Claire  Kinlaw  and  David  Harry 
Institute  of  Forest  Genetics,  USD  A,  Forest  Service 
Berkeley,  CA  94701 


Forest  trees  and  their  molecular 
genetics  were  the  focus  of  a recent 
conference  (Prouts  Neck,  Maine, 
USA,  May  20-23,  1994)  organized  by 
Michael  Greenwood  and  Keith 
Hutchison  (University  of  Maine, 
USA).  A group  of  70  researchers 
gathered  to  discuss  progress  toward 
understanding  and  manipulating 
molecular  processes  within  this 
diverse  group  of  economically  and 
ecologically  important  species. 

The  biology  of  forest  trees 
provides  both  distinct  challenges  and 
unique  opportunities.  Compared  to 
previous  meetings  of  this  research 
community,  significant  progress  has 
been  made  in  several  areas.  Research 
teams  continue  to  isolate  and  charac- 
terize new  genes,  while  transgenic 
plants,  especially  in  Populus,  are 
increasingly  being  used  to  study 
gene  function  in  vivo.  Genome 
mapping  has  also  matured.  In  earlier 
meetings,  presentations  described 
the  construction  of  genetic  linkage 
maps,  while  at  this  meeting  maps 
were  presented  as  tools  to  identify 
and  dissect  quantitative  trait  loci 
(QTL). 


Advantages  of  the  haploid 
genetics  offered  by  conifer  gameto- 
phytes  continue  to  be  exploited  for 
mapping  work  and  for  population 
surveys.  Recent  advances  in  model 
organisms  continue  to  influence 
studies  in  forest  trees.  Homeotic 
genes,  for  example,  are  being  sought 
for  flower  and  cone  development. 
Other  research  focuses  on  processes 
that  either  are  unique  to  trees  or  are 
simply  more  important  in  trees  than 
in  other  organisms.  For  example, 
lignin  and  its  related  biosynthetic 
pathways  are  affected  by  wounding 
and  stress  in  trees  as  in  other  plants, 
while  in  trees  alone,  lignin  also  is  a 
major  component  of  wood. 

Gene  Expression  Patterns 

As  highlighted  by  Olof  Olsson 
(Swedish  University  of  Agricultural 
Sciences,  Sweden),  woody  an- 
giosperms  show  fluctuations  in  gene 
expression  during  annual  cycles  of 
quiescence  and  "reesence"  in  addi- 
tion to  changes  observed  during 
development  and  in  response  to 
environmental  stresses.  With  the  goal 
of  increasing  pulp  production 


efficiency,  Wout  Boerjan's  laboratory 
(Universiteit  Gent,  Belgium)  has 
produced  transgenic  Populus  plants 
containing  antisense  constructs  for 
the  lignin  biosynthetic  enzymes  O- 
methyltransferase  (OMT)  and 
cinnamyl  alcohol  dehydrogenase 
(CAD).  Work  on  conifer  lignin  and 
its  role  in  development  continues 
despite  being  hindered  by  the  lack  of 
reliable  transformation  and  regenera- 
tion methods.  Working  on  Pinus 
taeda,  John  Mackay  (North  Carolina 
State  Biotechnology  Group,  USA, 
directed  by  Ron  Sederoff)  described 
the  isolation  and  characterization  of 
several  genes  encoding  lignin  biosyn- 
thetic enzymes  including  phenylala- 
nine ammonia  lyase  (PAL),  cinnamyl 
alcohol  dehydrogenase  (CAD),  and 
4-coumarate:CoA  ligase  (4-CL). 

A slightly  different  tack  toward 
understanding  xylogenesis  is  being 
taken  by  Mackay's  colleague, 
Malcolm  Campbell.  Because  myb- 
like  genes  play  an  important  role  in 
signal  transduction  pathways  in 
other  organisms,  such  genes  may 
also  control  conifer  xylem  develop- 
ment. Campbell  has  initiated  cloning 
of  Pinus  taeda  homologues.  A similar 
rationale  underlies  the  strategy  of 
Sharon  Regan  and  Bob  Rutledge 
(Petawawa  National  Forestry  Insti- 
tute, Canada)  in  their  efforts  to 
characterize  MADS  box  homeotic 
genes  controlling  cone  development 


38 


Probe 


Volume  4,  No.1/2 


in  Picea  mariana. 

With  the  goal  of  understanding 
the  role  of  flavonoids  in  root  forma- 
tion, Lise  Jouanin's  laboratory 
(INRA,  Versailles  Cedex,  France)  has 
produced  transgenic  Populus  and 
Juglans  containing  altered  levels  of 
chalcone  synthase  (CHS).  Carmen 
Diaz-Sala,  Keith  Hutchison,  and 
Mike  Greenwood  (University  of 
Maine,  USA)  are  investigating  the 
cellular  and  molecular  changes 
associated  with  the  organization  of 
root  primordia  in  Pinus  taeda.  In 
particular,  they  are  addressing  how 
the  cytoskeleton  orients  the  plane  of 
cellular  divisions  and  nuclear  reorga- 
nization. 

Engineering  for  pest  resistance 
using  proteinase  inhibitors  is  being 
done  by  several  groups  including 
those  of  Ned  Klopfenstein  (USDA-FS 
Center  for  Semiarid  Agroforestry, 
USA),  Lise  Jouanin,  and  Seguin 
Armand  (Universite  Laval,  Canada). 
Jouanin  has  found  a cysteine  pro- 
tease inhibitor  to  be  particularly 
effective  against  pests  which  contain 
high  levels  of  cysteine  proteases. 

Genes  responding  to  major 
environmental  stresses  have  been 
identified  in  stress-specific  cDNA 
libraries.  Genes  induced  by  drought 
stress  in  Pinus  taeda  (Shujun  Chang  et 
al.,  Texas  A&M  University,  USA) 
include  caffeoyl  CoA,  SAM  syn- 
thetase, chitinase,  and  a protein 
similar  to  an  animal  skin  matrix 
component.  Genes  induced  by  ozone 
(Dieter  Ernst,  Institut  fur 
Biocheniische  Pflanzenpathologie, 
Germany)  in  Pinus  sylvestris  include 
CAD,  stilbene  synthase, 
hydroxymethylglutaryl-CoA- 
synthase,  and  polyubiquitin.  As 


found  in  angiosperms,  certain  conifer 
genes  appear  to  be  induced  by  a 
number  of  environmental  stresses. 
For  example  chitinase  is  induced  by 
wounding,  fungal  infection,  and 
drought  (Haiguo  Wu,  Craig  Echt, 
and  John  Davis,  University  of 
Florida,  USA). 

To  expand  upon  the  identifica- 
tion of  new  conifer  genes,  Claire 
Kinlaw  (Institute  of  Forest  Genetics, 
USA)  has  initiated  "single-pass" 
sequencing  efforts  of  Pinus  taeda 
seedling  cDNAs  used  as  markers  by 
David  Neale  and  co-workers  (Insti- 
tute of  Forest  Genetics,  USA)  for 
genetic  maps.  These  identified 
sequences  will  provide  molecular 
tools  for  studying  conifer  genome 
organization  and  evolution.  Early 
results  have  been  encouraging  in  that 
a variety  of  genes  have  been  identi- 
fied including  those  encoding 
photosynthetic  proteins,  translation 
factors,  glycolytic  enzymes,  and 
stress-response  proteins. 

With  a systemic  point  of  view, 
Gary  Coleman  (Oregon  State  Univer- 
sity, USA)  proposed  a model  to 
explain  how  trees  regulate  autumn 
nitrogen  storage  and  spring 
remobilization  in  response  to  nitro- 
gen availability  and  photoperiod.  In 
this  model,  bark  and  leaves  commu- 
nicate with  each  other  using  a bark 
storage  protein  (BSP)  and  a leaf 
protein  encoded  by  Win4.  During 
short  days  or  high  levels  of  nitrogen, 
BSP  accumulates  in  bark  parenchyma 
while  the  Win4-encoded  protein  is 
repressed.  During  long  day  shoot 
growth,  BSP  is  degraded  while  the 
Win4-encoded  protein  accumulates. 


Genetic  Maps  and 
Quantitative  Trait  Loci 

Genetic  maps  using  two  alternative 
approaches  are  being  used  to  identify 
QTLs.  Lively  discussions  of  the 
merits  and  disadvantages  of  these 
two  alternate  approaches  accompa- 
nied formal  presentations.  Mitch 
Sewell  (Institute  of  Forest  Genetics, 
USA)  described  the  integration  of 
restriction  fragment  length  polymor- 
phism (RFLP)-based  linkage  maps 
from  two  Pinus  taeda  pedigrees.  This 
work  will  further  efforts  by  Neale 
and  his  co-workers  to  dissect  wood 
quality  traits  and  to  understand 
conifer  genome  organization  and 
evolution.  Several  members  of  the 
Forest  Biotechnology  Group  at  North 
Carolina  State  University  (USA) 
presented  RAPD-based  maps  includ- 
ing those  from  Eucalyptus  (Dario 
Grattapaglia)  for  the  identification  of 
QTL  controlling  sprouting  and 
rooting. 

New  Markers 

Several  laboratories  are  exploring  the 
use  of  length  polymorphism  among 
simple  sequence  repeats  (SSRs). 

Craig  Echt,  (USDA  Forest  Service, 
Rhinelander,  USA)  has  observed  that 
approximately  0.7%  of  the  Pinus 
strobus  genome  is  comprised  of  SSRs. 
Of  the  primer  pairs  tested  from  the 
flanking  sequences  of  Pinus  strobus 
SSR  loci,  approximately  65%  reliably 
amplify  DNA.  A high  proportion 
show  size  polymorphisms,  and  a 
significant  number  amplify  DNA 
from  other  conifer  taxa.  In  apparent 
contrast  to  these  observations,  Keith 
Hutchison  (University  of  Maine, 

USA)  has  observed  a low  level  of  size 
polymorphisms  among  SSR  alleles  in 


July  1993  -July  1994 


Probe 


39 


Larix  laricina. 

With  a similar  goal  of  develop- 
ing co-dominant  PCR-based  markers, 
but  using  a different  approach, 

David  Harry  (Institute  of  Forest 
Genetics,  USA)  is  designing  and 
testing  primers  based  upon  se- 
quences of  specific  Pinus  tnedn 
cDNAs.  Approximately  75%  of  the 
primer  pairs  reliably  amplify  ge- 
nomic DNA,  with  a high  proportion 
revealing  Mendelian  polymorphisms 
following  restriction  enzyme  diges- 
tion. Some  primers  amplify  only 
hard  pines,  others  amplify  all  pines, 
and  still  others  amplify  DNA  from 
other  conifer  taxa.  Hisato  Okuizumi 
presented  an  application  of  restric- 
tion landmark  genomic  scanning 
(RLGS)  to  large  genomes  by  includ- 
ing a restriction  trapper.  High-speed 
scanning  of  entire  genomes  and  the 
construction  of  genetic  maps  of 
individual  trees  from  a single  run 
with  several  hundred  loci  are  made 
possible.  As  an  example,  a profile  of 
Pinus  koraiensis  was  shown. 

Describing  Genome  Flux  and 
Evolution 

Because  seed  plants  represent  an 
ancient  lineage,  and  because  woody 
plants  have  long  generation  times, 
mechanisms  of  genetic  mutation  and 
genome  evolution,  as  well  as  rates  of 
species  evolution,  continue  to  be 
important  areas  of  study.  In  seeming 
contrast  to  low  levels  of  observed 
SSR  polymorphism  in  Larix  laricina, 
Hutchison  and  coworkers  have 
found  relatively  high  levels  of 
sequence  variation  within  genomic 
regions  encoding  proteins. 

In  addition,  they  have  observed 
segregation  distortion  of  alleles 


under  different  environments.  The 
apparent  contrast  between  the  high 
levels  of  polymorphism  among 
coding  regions  and  low  levels  of 
polymorphism  among  SSR  regions 
may  indicate  that  conifers  have  a 
relatively  efficient  mismatch  repair 
mechanism  and  may  thus  partially 
account  for  the  stability  of  conifer 
karyotypes. 

Jean  Bousquet  (Universite 
Laval,  Canada)  and  his  co-workers 
are  investigating  ancient  events 
during  the  evolution  of  seed  plants. 
After  carefully  calibrating  a molecu- 
lar clock,  they  established  that 
modern  gymnosperms  derived  from 
a single  lineage,  and  they  estimated 
divergence  times  to  have  occurred  as 
follows: 

liverworts  from  vascular  plants 
440  mya 

ferns  from  seed  bearing  plants 
400  mya 

flowering  from  cone  bearing  plants 
290  mya 

monocots  from  dicots 
200  mya 

Pinus  from  Pseudotsuga 
140  mya 

Ross  Whetten  (North  Carolina 
State  University,  USA)  is  exploiting 
the  idea  that  a tree's  crown  repre- 
sents a common  lineage  of  shoots 
with  known  separation  times.  Using 
visible  phenotypes  in  peach,  Whetten 
estimated  the  somatic  excision  rate  of 
a transposable  element.  The  rate  is 
relatively  higher  than  rates  reported 
for  annual  species  and  varies  among 
meristematic  layers.  This  notion  that 
different  shoots  within  the  crown  of 
a tree  can  be  genetically  distinct 
might  help  explain  how  long-lived 
trees  endure  pathogens  and  insect 
pests  with  shorter  generation  times. 


Genetic  Diversity  and 
Population  Structure 

In  addition  to  their  use  in  mapping, 
RAPDs  have  been  used  by  a number 
of  laboratories  for  estimating  genetic 
diversity  and  describing  population 
structure.  Natalie  Isabel  (Universite 
Laval,  Canada)  compared  estimates 
of  genetic  variation  within  and 
among  populations  of  Picea  mariana 
using  RAPDs  and  allozymes.  Results 
from  these  two  types  of  markers 
were  similar.  Linda  DeVerno 
(Petawawa  National  Forestry  Insti- 
tute, Canada)  surveyed  Pinus  resinosa 
using  400  RAPD  primers  and  found 
no  polymorphism.  Again  this  data 
supports  earlier  conclusions  based 
upon  allozymes. 

1995  Meeting 

The  next  tree  molecular  geneticists 
meeting  will  take  place  at 
Universiteit  Gent,  Belgium,  in 
combination  with  the  IUFRO  Somatic 
Cell  Genetics  Working  Party.  Those 
wishing  more  information  are 
encouraged  to  contact  Wout  Boerjan 
(Boerjan%research% 
RUG.genetica@genwetl.rug.ac.be). 

♦ 


MEMO 


Dear  Readers: 

Your  interests  are  impor- 
tant to  us.  Articles  and 
news  items  are  welcome. 

Next  deadline:  10/31/94 
Editor 


Probe 


Volume  4,  No.1/2 


Plant  Genome  Publications 


The  following  publications  are  available.  If  you  would 
like  to  receive  a copy,  check  off  the  title  and  mail  your 
request  to: 

Plant  Genome  Data  and  Information  Center 
National  Agricultural  Library,  USDA 
10301  Baltimore  Blvd.,  4th  Floor 
Beltsville,  MD  20705-2351 

CRIS/ICAR  Projects  and  Bibliographies: 

The  sponsored  research  projects  were  obtained  by 
searching  CRIS/ICAR  (USDA  and  CARC,  North 
America)  using  AGRISEARCH  Silver  Platter  CD. 
Bibliographies  were  obtained  by  searching  AGRICOLA 
Silver  Platter  CD. 

Application  of  RFLP  and  RAPD  Technologies  in 
Plant  Breeding.  January  1994.  Compiled  by  Andrew 
Kalinski. 

Application  of  PCR  Technology  to  Plant  Breeding. 
January  1994.  Compiled  by  Andrew  Kalinski. 

Patents  on  Transgenic  Plants.  February  1994. 
Compiled  by  Andrew  Kalinski. 


Plant  Genome  Data 
and  Information  Center 
USDA  - NAL 
10301  Baltimore  Blvd. 

Beltsville,  Maryland  20705-2351 


Soybean  Genome  Mapping.  January  1994.  Compiled 
by  Andrew  Kalinski. 

Tomato  and  Potato  Genome  Mapping.  January 
1994.  Compiled  by  Andrew  Kalinski. 

Transgenic  Maize.  February  1994.  Compiled  by 
Andrew  Kalinski. 

Transgenic  Soybean.  February  1994.  Compiled  by 
Andrew  Kalinski. 

Transgenic  Tomato.  February  1994.  Compiled  by 
Andrew  Kalinski. 


Miscellaneous  Publications: 

Prepared  by  the  Biotechnology  Information  Center  staff. 
AG/Biotechnology  Electronic  Information. 

ALF  (Agricultural  Library  Forum):  The  National 
Agricultural  Library's  Electronic  Bulletin  Board  Sys- 
tem - Brief  Guide.  Prepared  by  Karl  Schneider 
Biotechnology  Directories. 

Databases  Pertaining  to  Biotechnology. 

Guide  to  Information  Sources  in  Biotechnology. 
Newsletters  Pertaining  to  Agricultural  Biotechnology. 


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